Name: evaluation of fluid overload by bioelectrical impedance vectorial analysis
Uploaded: 2022-08-17T14:40:00
Description: Watch this Scientific Journal Video about Evaluation of Fluid Overload by Bioelectrical Impedance Vectorial Analysis at JoVE.com

The authors would like to thank Prof(s). Piccoli and Pastori of the Department of Medical and Surgical Sciences, University of Padova, Italy, for providing the BIVA software. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

The following protocol was approved (REF. 3057) and follows the guidelines of the human research ethics committee of Instituto Nacional de Ciencias M&#233;dicas y Nutrici&#243;n SZ. Furthermore, prior consent was obtained from the patients for this study.
NOTE: This procedure is to be used for measuring bioelectrical impedance analysis using tetrapolar multi-frequency equipment (see Table of Materials) and will provide accurate resistance and reactance values at a single frequ...

<ol>
	<li>da Silva, A. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Association+of+hyperhydration+evaluated+by+bioelectrical+impedance+analysis+and+mortality+in+patients+with+different+medical+conditions:+Systematic+review+and+meta-analyses.+Clinical+Nutrition+ASPEN+Association+of+hyperhydration+evaluated+by+bioelectrical.">Association of hyperhydration evaluated by bioelectrical impedance analysis and mortality in patients with different medical conditions: Systematic review and meta-analyses. Clinical Nutrition ASPEN Association of hyperhydration evaluated by bioelectrical.</a> Clinical Nutrition ESPEN. 28, 12-20 (2018).</li><li>Kammar-Garc&#237;a, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+Bioelectrical+Impedance+Analysis+parameters+for+the+detection+of+fluid+overload+in+the+prediction+of+mortality+in+patients+admitted+at+the+emergency+department.">Comparison of Bioelectrical Impedance Analysis parameters for the detection of fluid overload in the prediction of mortality in patients admitted at the emergency department.</a> Journal of Parenteral and Enteral Nutrition. 45 (2), 414-422 (2021).</li><li>Kammar-Garc&#237;a, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SOFA+score+plus+impedance+ratio+predicts+mortality+in+critically+ill+patients+admitted+to+the+emergency+department:+Retrospective+observational+study.">SOFA score plus impedance ratio predicts mortality in critically ill patients admitted to the emergency department: Retrospective observational study.</a> Healthcare (Basel). 10 (5), 810 (2022).</li><li>Frank Peacock, W., Soto, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+technique+of+fluid+status+assessment).">Current technique of fluid status assessment).</a> Congestive Heart Failure. 12, 45-51 (2010).</li><li>Lukaski, H. C., Vega-Diaz, N., Talluri, A., Nescolarde, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classification+of+hydration+in+clinical+conditions:+Indirect+and+direct+approaches+using+bioimpedance.">Classification of hydration in clinical conditions: Indirect and direct approaches using bioimpedance.</a> Nutrients. 11 (4), 809 (2019).</li><li>Bernal-Ceballos, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioimpedance+vector+analysis+in+stable+chronic+heart+failure+patients:+Level+of+agreement+single+and+multiple+frequency+devices.">Bioimpedance vector analysis in stable chronic heart failure patients: Level of agreement single and multiple frequency devices.</a> Clinical Nutrition ESPEN. 43, 206-211 (2021).</li><li>Uszko-Lencer, N. H., Bothmer, F., van Pol, P. E., Schols, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measuring+body+composition+in+chronic+heart+failure:+a+comparison+of+methods.">Measuring body composition in chronic heart failure: a comparison of methods.</a> European Journal of Heart Failure. 8 (2), 208-214 (2006).</li><li>Lukaski, H. C., Kyle, U. G., Kondrup, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+adult+malnutrition+and+prognosis+with+bioelectrical+impedance+analysis:+phase+angle+and+impedance+ratio.">Assessment of adult malnutrition and prognosis with bioelectrical impedance analysis: phase angle and impedance ratio.</a> Current Opinion in Clinical Nutrition and Metabolic. 20 (5), 330-339 (2017).</li><li>Piccoli, A., Rossi, B., Pillon, L., Bucciante, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+method+for+monitoring+body+fluid+variation+by+bioimpedance+analysis:+the+RXc+graph.">A new method for monitoring body fluid variation by bioimpedance analysis: the RXc graph.</a> Kidney International. 46 (2), 534-539 (1994).</li><li>Lukaski, H. C., Piccoli, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectrical+Impedance+Vector+Analysis+for+Assessment+of+Hydration+in+Physiological+States+and+Clinical+Conditions.">Bioelectrical Impedance Vector Analysis for Assessment of Hydration in Physiological States and Clinical Conditions.</a> Handbook of Anthropometry. , 287-305 (2012).</li><li>Piccoli, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bivariate+normal+values+of+the+bioelectrical+impedance+vector+in+adult+and+elderly+populations.">Bivariate normal values of the bioelectrical impedance vector in adult and elderly populations.</a> The American Journal of Clinical Nutrition. 61 (2), 269-270 (1995).</li><li>Roubenoff, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+bioelectrical+impedance+analysis+to+elderly+populations.">Application of bioelectrical impedance analysis to elderly populations.</a> The Journals of Gerontology. Series A, Biological Sciences and Medical Sciences. 52 (3), 129-136 (1997).</li><li>Espinosa-Cuevas, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bio+impedance+vector+an&#225;lisis+for+body+composition+in+Mexican+population.">Bio impedance vector an&#225;lisis for body composition in Mexican population.</a> Revista de Investigaci&#243;n Cl&#237;nica. 59 (1), 15-24 (2007).</li><li>Demirci, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impedance+ratio:+a+novel+marker+and+a+power+predictor+of+mortality+in+hemodialysis+patients.">Impedance ratio: a novel marker and a power predictor of mortality in hemodialysis patients.</a> International Urology and Nephrology. 48 (7), 1155-1162 (2016).</li><li>Plank, L. D., Li, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioimpedance+illness+marker+compared+to+phase+angle+as+a+predictor+of+malnutrition+in+hospitalized+patients.">Bioimpedance illness marker compared to phase angle as a predictor of malnutrition in hospitalized patients.</a> Clinical Nutrition. 32, 85 (2013).</li><li>Castillo-Martinez, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectrical+impedance+and+strength+measurements+in+patients+with+heart+failure:+comparison+with+functional+class.">Bioelectrical impedance and strength measurements in patients with heart failure: comparison with functional class.</a> Nutrition. 23 (5), 412-418 (2007).</li><li>Earthman, C. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Body+composition+tools+for+assessment+of+adult+malnutrition+at+the+bedside:+A+tutorial+on+research+considerations+and+clinical+applications.">Body composition tools for assessment of adult malnutrition at the bedside: A tutorial on research considerations and clinical applications.</a> Journal of Parenteral and Enteral Nutrition. 39 (7), 787-822 (2015).</li><li>Piccoli, A., Pastori, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=BIVA+software.">BIVA software.</a> Department of Medical and Surgical Sciences. , (2002).</li><li>Basso, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluid+management+in+the+intensive+care+unit:+bioelectrical+impedance+vector+analysis+as+a+tool+to+assess+hydration+status+and+optimal+fluid.">Fluid management in the intensive care unit: bioelectrical impedance vector analysis as a tool to assess hydration status and optimal fluid.</a> Blood Purification. 36 (3-4), 192-199 (2013).</li><li>Piccoli, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectrical+impedance+measurement+for+fluid+status+assessment.">Bioelectrical impedance measurement for fluid status assessment.</a> Contributions to Nephrology. 164, 143-152 (2010).</li><li>National Institutes of Health Technology. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectrical+impedance+analysis+in+body+composition+measurement:+National+Institutes+of+Health+Technology+Assessment+Conference+Statement.">Bioelectrical impedance analysis in body composition measurement: National Institutes of Health Technology Assessment Conference Statement.</a> The American Journal of Clinical Nutrition. 64, 524-532 (1996).</li><li>Silva, A. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lack+of+agreement+of+in+vivo+raw+bioimpedance+measurements+obtained+from+two+single+and+multifrequency+bioelectrical+impedance+devices.">Lack of agreement of in vivo raw bioimpedance measurements obtained from two single and multifrequency bioelectrical impedance devices.</a> European Journal of Clinical Nutrition. 73 (7), 1077-1083 (2019).</li><li>Mulasi, U., Kuchnia, A. J., Cole, A. J., Earthman, C. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioimpedance+at+the+bedside:+current+applications,+limitations,+and+opportunities.">Bioimpedance at the bedside: current applications, limitations, and opportunities.</a> Nutrition in Clinical Practice. 30 (2), 180-193 (2015).</li><li>Chabin, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioimpedance+analysis+is+safe+in+patients+with+implanted+cardiac+electronic+devices.">Bioimpedance analysis is safe in patients with implanted cardiac electronic devices.</a> Clinical Nutrition. 38 (2), 806-811 (2019).</li><li>Gonz&#225;lez-Correa, C. H., Caicedo-Eraso, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectrical+impedance+analysis+(BIA):+a+proposal+for+standardization+of+the+classical+method+in+adults.">Bioelectrical impedance analysis (BIA): a proposal for standardization of the classical method in adults.</a> Journal of Physics: Conference Series. 47, 407 (2012).</li><li>Di Somma, S., Gori, C. S., Grandi, T., Risicato, M. G., Salvatori, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluid+assessment+and+management+in+the+emergency+department.">Fluid assessment and management in the emergency department.</a> Contributions to Nephrology. 164, 227-236 (2010).</li><li>Kammar-Garc&#237;a, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mortality+in+adult+patients+with+fluid+overload+evaluated+by+BIVA+upon+admission+to+the+emergency+department.">Mortality in adult patients with fluid overload evaluated by BIVA upon admission to the emergency department.</a> Postgraduate Medical Journal. 94 (1113), 386-391 (2018).</li></ol>

It is important to mention that different bioelectrical impedance analysis (BIA) approaches have been proposed in the published literature, including the use of multiple frequencies at 1-500 kHz (MF-BIA), phase-sensitive single frequency (SF-BIA) at 50 kHz, and spectroscopic BIA at 5 kHz to 2 MHz. Studies have provided inconsistent results, concerning the agreement regarding single- and multiple-frequency BIA equipment6, including source current, frequency, total impedance range over which the cur...

Fluid overload (FO), defined as an excess of total body fluid or a relative excess in one or more fluid compartments1, is frequently observed in critically ill patients and is associated with higher morbidity and mortality1,2,3. The range of alterations in hydration status is wide; can indicate renal, cardiac, or hepatic failure; and/or maybe the result of excessive oral intake or iatrogenic error4. Routine assessment of hydration status is challenging in emergency departments, as the gold standard of radioisotopic volume measurement requires specialized techniques, is costly and time-consuming, and may fail to identify early disturbances in hydration status. Hence, other less-accurate methods are generally used, including clinical examination and accumulated fluid balance (volume in mL in 24 h)5. Accurate and sensitive determination of fluid volume status is necessary to help clinicians in controlling body fluids, managing intravenous fluid administration, and maintaining hemodynamic stability, thus allowing patients to receive early treatment3,5,6. Errors in the volume assessment can lead to a lack of necessary treatment or to implementation of unnecessary therapy, such as excess fluid administration, both of which are related to increased hospitalization costs, complications, and mortality4.
Interest has recently increased in bioelectrical impedance analysis (BIA), which has been considered an alternative method for the classification of an individual&#39;s hydration status. BIA is a safe, non-invasive, portable, quick, bed-side, and easy-to-use method, designed for the estimation of body compartment composition. The analysis measures the opposition generated by soft tissues to the flow of an injected alternating electric current into the body (800 &#181;A), through four surface electrodes placed on the hands and feet. Total body water estimated by BIA has been shown to have a high correlation with that obtained by deuterium dilution (r = 0.93, p = 0.01)7.
Phase-sensitive BIA devices evaluate the direct measurement of phase angle and impedance (Z50), obtaining the resistance (R) and reactance (Xc) in single-frequency mode (50 kHz) or multi-frequency mode (5 kHz to 200 kHz)8. Dividing the R and Xc values by the subject&#39;s height (in m) squared-to control for inter-individual differences in conductor length-and plotting them in an R-Xc graph is the method used in bioelectrical impedance vector analysis (BIVA) to estimate the fluid status. BIVA is an alternative impedance approach, developed by Piccoli et al.9, which uses the spatial relationship between R (i.e., the opposition to the flow of an alternating current through intra- and extra-cellular ionic solutions) and Xc to assess soft tissue hydration, independent of the multiple-regression prediction equations generated in limited and specific samples10. Therefore, the classification of fluid status is more precise and accurate than the quantification of total body water. The R and Xc values of a subject produce a vector whose position can be evaluated relative to tolerance intervals in the R-Xc graph, which can be interpreted as indicating normal or abnormal hydration, based on the distance from the mean vector derived from a healthy reference population11,12,13.
In a previous study, we compared different bioelectrical impedance analysis parameters for the detection of fluid overload and prediction of mortality in patients admitted to an emergency department (ED) and demonstrated that BIVA (relative risk = 6.4; 95% confidence interval from 1.5 to 27.9; p = 0.01) and impedance ratio (relative risk = 2.7; 95% confidence interval from 1.1 to 7.1; p = 0.04) improved the estimation of the probability of 30-day mortality3.
Fluid overload can also be estimated using the impedance ratio (imp-R), which is the ratio between impedance measured at 200 kHz and impedance measured at 5 kHz obtained by the multi-frequency bioelectrical impedance equipment. Imp-R considers conduction in total body water (Z200) and in extracellular water fluid spaces (Z5). The penetration of a current into cells is frequency-dependent and, the 200/5 kHz ratio describes the ratio of greater to lesser current entry into cells3,8. If the difference between these two values decreases over time, it may indicate that the cells are becoming less healthy14.
Imp-R values &#8804;0.78 in males and &#8804;0.82 in females have been observed in healthy individuals15. Values nearer to 1.0 indicate that the two impedances are closer to each other, and the body cell is less healthy. In the case of critical illness, the resistance of the cell membrane at 5 kHz is reduced, and the difference between the impedance values at 5 and 200 kHz is markedly lower, indicating cellular worsening3. Values &#62; 1.0 suggest device error16,17. Thus, the objective of the present study is to demonstrate how to evaluate the presence of fluid overload through bioelectrical impedance vectorial analysis, as well as by using the impedance ratio, measured with tetrapolar multi-frequency equipment in patients admitted to the emergency department.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Alcohol 70% swabs</td>
			<td>NA</td>
			<td>NA</td>
			<td>Any brand can be used</td>
		</tr>
		<tr>
			<td>BIVA software 2002</td>
			<td>NA</td>
			<td>NA</td>
			<td>Is a sofware created for academic use, can be download in http://www.renalgate.it/formule_calcolatori/bioimpedenza.htm in &#34;LE FORMULE DEL Prof. Piccoli&#34; section</td>
		</tr>
		<tr>
			<td>Chlorhexidine Wipes</td>
			<td>NA</td>
			<td>NA</td>
			<td>Any brand can be used</td>
		</tr>
		<tr>
			<td>Examination table</td>
			<td>NA</td>
			<td>NA</td>
			<td>Any brand can be used</td>
		</tr>
		<tr>
			<td>Leadwires square socket</td>
			<td>BodyStat</td>
			<td>SQ-WIRES</td>
			<td></td>
		</tr>
		<tr>
			<td>Long Bodystat 0525 electrodes</td>
			<td>BodyStat</td>
			<td>BS-EL4000</td>
			<td></td>
		</tr>
		<tr>
			<td>Quadscan 4000 equipment</td>
			<td>BodyStat</td>
			<td>BS-4000</td>
			<td>Impedance measuring range: 20 - 1300 &#937; ohms 
			Test Current: 620 &#956;A 
			Frequency: 5, 50, 100, 200 kHz 
			Accuracy: Impedance 5 kHz: +/- 2 &#937; 
			Impedance 50 kHz: +/- 2 &#937; 
			Impedance 100 kHz: +/- 3 &#937; 
			Impedance 200 kHz: +/- 3 &#937; 
			Resistance 50 kHz: +/- 2 &#937; 
			Reactance 50 kHz: +/- 1 &#937; 
			Phase Angle 50 kHz: +/- 0.2&#176; 
			Calibration: A resistor is supplied for independent verification from time to time. The impedance value should read between 496 and 503 &#937;.</td>
		</tr>
	</tbody>
</table>

evaluation of fluid overload by bioelectrical impedance vectorial analysis

Early detection and management of fluid overload are critically important in acute illness, as the impact of therapeutic intervention can result in decreased or increased mortality rates. Accurate fluid status assessment entails appropriate therapy. Unfortunately, as the gold standard method of radioisotopic fluid measurement is costly, time-consuming, and lacks sensitivity in the acute care clinical setting, other less-accurate methods are typically used, such as clinical examination or 24 h output. Bioelectrical impedance vectorial analysis (BIVA) is an alternative impedance-based approach, where the raw parameter resistance and reactance of a subject are plotted to produce a vector, the position of which can be evaluated relative to tolerance intervals in an R-Xc graph. The fluid status is then interpreted as normal or abnormal, based on the distance from the mean vector derived from a healthy reference population. The objective of the present study is to demonstrate how to evaluate the presence of fluid overload through bioelectrical impedance vectorial analysis and the impedance ratio measured with tetrapolar multi-frequency equipment in patients admitted to the emergency department.

As an example of the method presented above, we present the results for two women admitted to the emergency department. Bioelectrical impedance analysis was assessed at admission using a phase-sensitive multi-frequency device (see Table of Materials), and the obtained resistance (R) and reactance (Xc) values were used to calculate the BIVA graph. The results show that patients with overhydration had worse prognoses and clinical characteristics such as SOFA and Charlson index scores, which are related to ...

Watch this Scientific Journal Video about Evaluation of Fluid Overload by Bioelectrical Impedance Vectorial Analysis at JoVE.com

Evaluation of Fluid Overload by Bioelectrical Impedance Vectorial Analysis

In this study, we demonstrate how to evaluate the presence of fluid overload through bioelectrical impedance vectorial analysis (BIVA) and the impedance ratio measured using tetrapolar multi-frequency equipment in patients admitted to the emergency department. BIVA and impedance ratio are reliable and useful tools to predict poor outcomes.

Early detection and management of fluid overload are critically important in acute illness, as the impact of therapeutic intervention can result in ...

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A Real-time Electrical Impedance Based Technique to Measure Invasion of Endothelial Cell Monolayer by Cancer Cells

Metastatic dissemination of malignant cells requires degradation of basement membrane, attachment of tumor cells to vascular endothelium, retraction of endothelial junctions and finally invasion and migration of tumor cells through the endothelial layer to enter the bloodstream as a means of transport to distant sites in the host1-3. Once in the circulatory system, cancer cells adhere to capillary walls and extravasate to the surrounding tissue to form metastatic tumors4,5. The various components of tumor cell-endothelial cell interaction can be replicated in vitro by challenging a monolayer of human umbilical vein endothelial cells (HUVEC) with cancer cells. Studies performed with electron and phase-contrast microscopy suggest that the in vitro sequence of events fairly represent the in vivo metastatic process6. Here, we describe an electrical-impedance based technique that monitors and quantifies in real-time the invasion of endothelial cells by malignant tumor cells.
Giaever and Keese first described a technique for measuring fluctuations in impedance when a population of cells grow on the surface of electrodes7,8. The xCELLigence instrument, manufactured by Roche, utilizes a similar technique to measure changes in electrical impedance as cells attach and spread in a culture dish covered with a gold microelectrode array that covers approximately 80% of the area on the bottom of a well. As cells attach and spread on the electrode surface, it leads to an increase in electrical impedance9-12. The impedance is displayed as a dimensionless parameter termed cell-index, which is directly proportional to the total area of tissue-culture well that is covered by cells. Hence, the cell-index can be used to monitor cell adhesion, spreading, morphology and cell density.
The invasion assay described in this article is based on changes in electrical impedance at the electrode/cell interphase, as a population of malignant cells invade through a HUVEC monolayer (Figure 1). The disruption of endothelial junctions, retraction of endothelial monolayer and replacement by tumor cells lead to large changes in impedance. These changes directly correlate with the invasive capacity of tumor cells, i.e., invasion by highly aggressive cells lead to large changes in cell impedance and vice versa. This technique provides a two-fold advantage over existing methods of measuring invasion, such as boyden chamber and matrigel assays: 1) the endothelial cell-tumor cell interaction more closely mimics the in vivo process, and 2) the data is obtained in real-time and is more easily quantifiable, as opposed to end-point analysis for other methods.

This article describes an in vitro technique for monitoring cancer cells invading through a monolayer of endothelial cells. The data is acquired in real-time as a function of changes in impedance on the surface of electrodes at the well bottom.

Metastatic dissemination of malignant cells requires degradation of basement membrane, attachment of tumor cells to vascular endothelium, retraction of ...

Mouse Model of Intraluminal MCAO: Cerebral Infarct Evaluation by Cresyl Violet Staining

Stroke is the third cause of mortality and the leading cause of disability in the World. Ischemic stroke accounts for approximately 80% of all strokes. However, the thrombolytic tissue plasminogen activator (tPA) is the only treatment of acute ischemic stroke that exists. This led researchers to develop several ischemic stroke models in a variety of species. Two major types of rodent models have been developed: models of global cerebral ischemia or focal cerebral ischemia. To mimic ischemic stroke in patients, in whom approximately 80% thrombotic or embolic strokes occur in the territory of the middle cerebral artery (MCA), the intraluminal middle cerebral artery occlusion (MCAO) model is quite relevant for stroke studies. This model was first developed in rats by Koizumi et al. in 1986 1. Because of the ease of genetic manipulation in mice, these models have also been developed in this species 2-3.
	Herein, we present the transient MCA occlusion procedure in C57/Bl6 mice. Previous studies have reported that physical properties of the occluder such as tip diameter, length, shape, and flexibility are critical for the reproducibility of the infarct volume 4. Herein, a commercial silicon coated monofilaments (Doccol Corporation) have been used. Another great advantage is that this monofilament reduces the risk to induce subarachnoid hemorrhages. Using the Zeiss stereo-microscope Stemi 2000, the silicon coated monofilament was introduced into the internal carotid artery (ICA) via a cut in the external carotid artery (ECA) until the monofilament occludes the base of the MCA. Blood flow was restored 1 hour later by removal of the monofilament to mimic the restoration of blood flow after lysis of a thromboembolic clot in humans. The extent of cerebral infarct may be evaluated first by a neurologic score and by the measurement of the infarct volume. Ischemic mice were thus analyzed for their neurologic score at different post-reperfusion times. To evaluate the infarct volume, staining with 2,3,5-triphenyltetrazolium chloride (TTC) was usually performed. Herein, we used cresyl violet staining since it offers the opportunity to test many critical markers by immunohistochemistry. In this video, we report the MCAO procedure; neurological scores and the evaluation of the infarct volume by cresyl violet staining.

The intraluminal middle cerebral occlusion model in mice is herein presented. The extent of cerebral infarct is evaluated by a neurologic score and cresyl violet staining, an alternative staining to TTC, offering the great advantage to test in parallel many interest markers.

Stroke is the third  cause of mortality and the leading cause of disability in the World. Ischemic  stroke accounts for approximately 80% of all strokes. ...

Evaluation of Patients' Posture and Gait Profile After Lumbar Fusion Surgery by Video Rasterstereography and Treadmill Gait Analysis

This protocol provides guidance on how to perform high resolution video rasterstereography and treadmill gait analysis on patients after lumbar fusion surgery to obtain results about altered variables of gait and posture. These observed changes can then be correlated with the patient-reported outcome measure of pain relief. The rasterstereographic device projects lines of parallel light onto the surface of the tested subject&#39;s back. The deformation of these lines is recognized by the device. From these data, a special software then generates a 3-D profile based on the principle of triangulation. With an inaccuracy of only 0.2 mm it can measure changes in posture at very high precision. Gait and stance parameters are recorded using a treadmill equipped with an electric sensor mat that contains 10,200 miniature force sensors in the registering zone under the belt. Initial walking speed on the treadmill is 0.5 km/h. Speed is then gradually increased by increments of 0.1 km/h until each subject reaches his or her individual maximum well tolerable walking speed. At this speed, parameters are recorded during a 20 s measurement interval. Subjects are tested barefoot and without holding a handrail. Among various other parameters, stride width, step length, stance phase and foot rotation are measured. Both methods used reportedly have a high intra- and inter-observer reliability. The advantage of these highly accurate techniques is that they offer an objective and very detailed perspective on changes in the patient&#39;s posture and gait. Due to the amount of data generated, these techniques are, however, not so much suitable for everyday routine use, but rather interesting to scientifically evaluate long term alterations in posture and gait in patients like for example after lumbar fusion surgery.

Evaluation of Patients&#039; Posture and Gait Profile After Lumbar Fusion Surgery by Video Rasterstereography and Treadmill Gait Analysis

Here, we present a protocol to analyze the posture and the gait of patients after lumbar fusion surgery by means of high-resolution video rasterstereography and a treadmill equipped with an integrated sensor mat. Allowing critical functional postoperative evaluation on a less subjective level may enhance accuracy and reliability of indication for surgery.

This protocol provides guidance on how to perform high resolution video rasterstereography and treadmill gait analysis on patients after lumbar fusion ...

Synovial Fluid Analysis to Identify Osteoarthritis

Synovial fluid (SF) analysis is important in diagnosing osteoarthritis (OA). Macroscopic and microscopic features, including total and differential white blood cell (WBC) count, help define the non-inflammatory nature of SF, which is a hallmark of OA. In patients with OA, WBC in SF samples usually does not exceed 2000 cells per microliter, and the percentage of inflammatory cells, such as neutrophils, is very low or absent. Calcium crystals are frequent in SF collected from OA patients. Although their role in the pathogenesis of OA remains unclear, they have been associated with a mild inflammatory process and a more severe disease progression. Recently, calcium crystals have been described in both the early and late stages of OA, indicating that they may play a vital role in diagnosing different clinical subsets of OA and pharmacological treatment. The overall goal of SF analysis in OA is two-fold: to ascertain the non-inflammatory degree of SF and to highlight the presence of calcium crystals.

Synovial fluid analysis under transmitted, polarized, and compensated light microscopy is used to evaluate the inflammatory or non-inflammatory nature of a sample through simple steps. It is particularly useful in osteoarthritis to detect calcium crystals and identify a more severe subset of osteoarthritis.

Synovial fluid (SF) analysis is important in diagnosing osteoarthritis (OA). Macroscopic and microscopic features, including total and differential white ...

Analysis of Raw and Processed Cyperi Rhizoma Samples Using Liquid Chromatography-Tandem Mass Spectrometry in Rats with Primary Dysmenorrhea

Cyperi rhizoma (CR) is widely used in gynecology and is a general medicine for treating women's diseases in China. Since the analgesic effect of CR is enhanced after processing with vinegar, CR processed with vinegar (CRV) is generally used clinically. However, the mechanism by which the analgesic effect is enhanced by vinegar processing is unclear. In this study, the ultra-high pressure liquid chromatographytandem mass spectrometry (UPLC-MS/MS) technique was used to examine changes in the blood levels of the exogenous constituents and metabolites between CR-treated and CRV-treated rats with dysmenorrhea. The results revealed differing levels of 15 constituents and two metabolites in the blood of these rats. Among them, the levels of (-)-myrtenol and [(1R,2S,3R,4R)-3-hydroxy-1,4,7,7-tetramethylbicyclo[2.2.1]hept-2-yl]acetic acid in the CRV group were considerably higher than in the CR group. CRV reduced the level of 2-series prostanoids and 4-series leukotrienes with proinflammatory, platelet aggregation, and vasoconstriction activities and provided analgesic effects by modulating arachidonic acid and linoleic acid metabolism and the biosynthesis of unsaturated fatty acids. This study revealed that vinegar processing enhances the analgesic effect of CR and contributes to our understanding of the mechanism of action of CRV.

Here, a comparative analysis of raw and processed Cyperi rhizoma (CR) samples is presented using ultra-high performance liquid chromatography-high-resolution tandem mass spectrometry (UPLC-MS/MS) in rats with primary dysmenorrhea. The changes in blood levels of the metabolites and the sample constituents were examined between rats treated with CR and CR processed with vinegar (CRV).

Cyperi rhizoma (CR) is widely used in gynecology and is a general medicine for treating women's diseases in China. Since the analgesic effect of CR is ...

Modified Arteriovenous Fistula Surgery to Establish the RV Volume Overload Mouse Model

Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV myocardium may respond differently to VO in children compared to adults. The present study aims to establish a postnatal RV VO model in mice using a modified abdominal arteriovenous fistula. To confirm the creation of VO and the following morphological and hemodynamic changes of the RV, abdominal ultrasound, echocardiography, and histochemical staining were performed for 3 months. As a result, the procedure in postnatal mice showed an acceptable survival and fistula success rate. In VO mice, the RV cavity was enlarged with a thickened free wall, and the stroke volume was increased by about 30%-40% within 2 months after surgery. Thereafter, the RV systolic pressure increased, corresponding pulmonary valve regurgitation was observed, and small pulmonary artery remodeling appeared. In conclusion, modified arteriovenous fistula (AVF) surgery is feasible to establish the RV VO model in postnatal mice. Considering the probability of fistula closure and elevated pulmonary artery resistance, abdominal ultrasound and echocardiography must be performed to confirm the model status before application.

Author Spotlight: Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model  

This protocol presents the establishment and confirmation of a postnatal right ventricular volume overload (VO) model in mice with abdominal arteriovenous fistula (AVF), which can be applied to investigate how VO contributes to postnatal heart development.

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV ...

Predicting Prognosis in Acute Heart Failure Using Phase Angle &amp; BIVA Z-Scores

Clinical Application of Phase Angle and BIVA Z-Score Analyses in Patients Admitted to an Emergency Department with Acute Heart Failure

Acute heart failure is characterized by neurohormonal activation, which leads to sodium and water retention and causes alterations in body composition, such as increased body fluid congestion or systemic congestion. This condition is one of the most common reasons for hospital admission and has been associated with poor outcomes. The phase angle indirectly measures intracellular status, cellular integrity, vitality, and the distribution of spaces between intracellular and extracellular body water. This parameter has been found to be a predictor of health status and an indicator of survival and other clinical outcomes. In addition, phase angle values of &lt;4.8&#176; upon admission were associated with higher mortality in patients with acute heart failure. However, low phase angle values may be due to alterations-such as the shifting of fluids from an intracellular body water (ICW) compartment to an ECW (extracellular body water) compartment and a concurrent decrease in body-cell mass (which can reflect malnutrition)-that are present in heart failure. Thus, a low phase angle may be due to overhydration and/or malnutrition. BIVA provides additional information about the body-cell mass and congestion status with a graphical vector (R-Xc graph). In addition, a BIVA Z-score analysis (the number of standard deviations from the mean value of the reference group) that has the same pattern as that of the ellipses for the percentiles on the original R-Xc graph can be used to detect changes in soft-tissue mass or tissue hydration and can help researchers compare changes in different study populations. This protocol explains how to obtain and interpret phase angle values and BIVA Z-score analyses, their clinical applicability, and their usefulness as a predictive marker for the prognosis of a 90-day event in patients admitted to an emergency department with acute heart failure.

Author Spotlight: Unveiling Prognostic Indicators in Heart Failure - The Role of Phase Angle and Bioelectrical Impedance Analysis 

In this protocol, we explain how to obtain and interpret phase angle values and bioelectrical impedance vectorial analysis (BIVA) Z-score obtained by bioelectrical impedance in patients with acute heart failure admitted to the Emergency Department and their clinical applicability as a predictive marker for the prognosis of a 90-day event.

Acute heart failure is characterized by neurohormonal activation, which leads to sodium and water retention and causes alterations in body composition, ...

Developing an Exercise Program for Rheumatoid Arthritis Patients

Evaluation of Changes in Hydration and Body Cell Mass with Bioelectrical Impedance Analysis after Exercise Program for Rheumatoid Arthritis Patients

Rheumatoid arthritis (RA) is a debilitating disease that can result in complications such as rheumatoid cachexia. While physical exercise has shown benefits for RA patients, its impact on hydration and body cell mass remains uncertain. The presence of pain, inflammation, and joint changes often restrict activity and make traditional body composition assessments unreliable due to altered hydration levels. Bioelectrical impedance is a commonly used method for estimating body composition, but it has limitations since it was primarily developed for the general population and does not consider changes in body composition. On the other hand, bioelectrical impedance vectorial analysis (BIVA) offers a more comprehensive approach. BIVA involves graphically interpreting resistance (R) and reactance (Xc), adjusted for height, to provide valuable information about hydration status and the integrity of the cell mass.
Twelve women with RA were included in this study. At the beginning of the study, hydration and body cell mass measurements were obtained using the BIVA method. Subsequently, the patients participated in a six-month dynamic exercise program encompassing cardiovascular capacity, strength, and coordination training. To evaluate changes in hydration and body cell mass, the differences in the R and Xc parameters, adjusted for height, were compared using BIVA confidence software. The results showed notable changes: resistance decreased after the exercise program, while reactance increased. BIVA, as a classification method, can effectively categorize patients into dehydration, overhydration, normal, athlete, thin, cachectic, and obese categories. This makes it a valuable tool for assessing RA patients, as it provides information independent of body weight or prediction equations. Overall, the implementation of BIVA in this study shed light on the effects of the exercise program on hydration and body cell mass in RA patients. Its advantages lie in its ability to provide comprehensive information and overcome the limitations of traditional body composition assessment methods.

Author Spotlight: Implementation of BIVA for Analyzing Disease Risk Factors in Patients with Low Body Cell Mass 

This protocol assesses alterations in hydration and body cell mass status using bioelectrical impedance vectorial analysis following a dynamic exercise program designed for patients with rheumatoid arthritis. The dynamic exercise program itself is detailed, highlighting its components focused on cardiovascular capacity, strength, and coordination. The protocol details steps, instruments, and limitations.

Rheumatoid arthritis (RA) is a debilitating disease that can result in complications such as rheumatoid cachexia. While physical exercise has shown ...

Evaluation of Hydration Status by Bioelectrical Impedance Vector Analysis in Patients with Ischemic Heart Disease Undergoing Exercise Stress Test

Ischemic heart disease (IHD) represents a group of clinical syndromes characterized by myocardial ischemia, leading to an impairment in the myocardial blood supply and compromised perfusion. Several clinical variables assessed through a stress test, such as oxygen uptake (VO2) and heart rate oxygen pulse (HR/O2), have been attributed as cardiopulmonary prognostic factors in patients with IHD. However, other factors like hydration status (HS), potentially affecting the cardiopulmonary response, have been barely addressed. Unbalanced HS has a short-term effect on plasma volume and the sympathetic nervous system, which impacts blood volume, and lowers VO2 and HR/O2. Recently, bioelectrical impedance analysis (BIA), a method based on the opposition of body tissues (including fluid volume) to a low electrical current, has been widely used to assess HS by obtaining two components: resistance (R) and reactance (Xc) and using prediction formulas. However, several limitations as chronic illness or abnormal fluid status, may affect the results. In this sense, alternative BIA methods, such as bioelectrical impedance vector analysis (BIVA), have become relevant. R and Xc (adjusted by height) result in a vector plotted on the R/Xc graph, which allows interpreting the HS as normal or abnormal according to the distance of the mean vector. This study aims to describe how to determine HS by BIVA using a single-frequency device and compare the results with the cardiopulmonary response in patients with IHD.

Hydration status imbalances can have a short-term effect on direct and indirect determinants of oxygen uptake and pulse, and morbidity and mortality prognostic factors in ischemic heart disease. This protocol describes the technique for assessment of hydration status through bioelectrical impedance vector analysis and cardiopulmonary response during exercise stress test.

Ischemic heart disease (IHD) represents a group of clinical syndromes characterized by myocardial ischemia, leading to an impairment in the myocardial ...

Analyzing Bioelectrical Impedance Analysis (BIA) Raw Parameters as a Predictive Marker for the Prognosis of Acute Heart Failure

Begin by removing the shoe and sock from the patient's right foot. Also remove all metal objects in contact with the patient's skin, such as bracelets, watches, rings, and chains. Position the patient's supine or semi-Fowler position with their legs and arms spread at approximately 45 degrees.To measure BIA, clean the surfaces using a 70%alcohol pad and allow them to dry completely before placing the electrodes. Download the BIVAtolerance software by Piccoli. Open the Reference populations sheet.Copy the appropriate reference population based on the patient's characteristics and paste it in the first yellow row. Click on the Z-score sheet. Insert the reference population and enter the medical record number of each patient into the Subject ID field.Next, input a number between one and 10 in the Group Code field. Insert the resistance value obtained with BIA and adjusted by height in meters into the resistance divided by height subject field and the reactance value into the reactance divided by height subject field. Then input one in the Drawing option field to generate a plot.Navigate to the spreadsheet program menu, select the Add-ins tab and click the Calculate button. Click on the Z-graph sheet within the spreadsheet program. Navigate to the Add-ins tab.Click the NEW GRAPH button. Then proceed with conducting BIVA Z-score and phase angle analyses.