74 Zukow Walery, Flyunt Igor-Severyn S., Ponomarenko Ruslan B., Rybak Nadiya Y., Fil’ Vitaliy M., Kovalchuk Halyna Y., Sarancha Serhiy M., Nahurna Yaryna V. Polivariant change of step-test under the influence of natural adaptogens and their accompaniments. Pedagogy and Psychology of Sport. 2020;6(2):74-84. elSSN 2450-6605. DOI http://dx.doi.org/10.12775/PPS.2020.06.02.007 https://apcz.umk.pl/czasopisma/index.php/PPS/article/view/PPS.2020.06.02.007 https://zenodo.org/record/3767282 The journal has had 5 points in Ministry of Science and Higher Education parametric evaluation. § 8. 2) and § 12. 1. 2) 22.02.2019. © The Authors 2020; This article is published with open access at Licensee Open Journal Systems of Nicolaus Copernicus University in Torun, Poland Open Access. This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author (s) and source are credited. This is an open access article licensed under the terms of the Creative Commons Attribution Non commercial license Share alike. (http://creativecommons.org/licenses/by-nc-sa/4.0/) which permits unrestricted, non commercial use, distribution and reproduction in any medium, provided the work is properly cited. The authors declare that there is no conflict of interests regarding the publication of this paper. Received: 05.04.2020. Revised: 16.04.2020. Accepted: 26.04.2020. Polivariant change of step-test under the influence of natural adaptogens and their accompaniments Walery Zukow1, Igor-Severyn S. Flyunt2, Ruslan B. Ponomarenko3, Nadiya Y. Rybak3, Vitaliy M. Fil’2, Halyna Y. Kovalchuk2, Serhiy M. Sarancha4, Yaryna V. Nahurna5 1Nicolaus Copernicus University, Torun, Poland w.zukow@wp.pl 2Ivan Franko Pedagogical University, Drohobych, Ukraine fillvitalij@gmail.com 3Communal Hospital, Truskavets’, Ukraine karapuzia@ukr.net 4Children’s Sanatorium “Dzherelo”, Truskavets’, Ukraine 5Danylo Halyts’kyǐ National Medical University, L’viv, Ukraine Corresponding Author Walery Zukow, E-mail: w.zukow@wp.pl Abstract Background. Typically, the step test is used to assess the physical performance that is determined by the condition of the cardio-respiratory system. The latter, in turn, is associated with the nervous, endocrine and immune systems, which collectively determine the level of adaptation of the body. The purpose of this study is to analyze the effect on the step test of natural adaptogens and associated changes in a number of parameters of cardio-respiratory, autonomic nervous and immune systems and metabolism. Material and methods. The target was 80 children of both sexes, ages 10-17, arriving at the clinical sanatorium "Dzherelo" of Truskavets spa from radionuclide contaminated territories. The main subject of the study was the Popovych’s step test. State of autonomous nervous and immune systems was assessed as well as http://dx.doi.org/10.12775/PPS.2020.06.02.007 https://apcz.umk.pl/czasopisma/index.php/PPS/article/view/PPS.2020.06.02.007 https://zenodo.org/record/3767282 mailto:w.zukow@.wp.pl mailto:fillvitalij@gmail.com mailto:karapuzia@ukr.net mailto:w.zukow@.wp.pl 75 others functional and metabolic tests. Results. The contingent surveyed was retrospectively divided into three qualitatively distinct groups. The largest group (53,7%) was the group of children with initially reduced or normal levels of Popovych’s Step Test Index (PSI), in which it increased. However, in 28,8% of children PSI remained stable at different initial levels, and in 17,5% even decreased, without falling below the lower limit of normal. Discriminant analysis was subsequently applied to identify constellations of parameters that change are characteristic of each cluster. The program selected as characteristic changes in 2 parameters of autonomous nervous system, 2 functional, 4 metabolic and 3 immune parameters as well as blood trombocytes level. The correct classification of the positive actotropic effect is 93,0%; neutral 65,2%; negative 64,3%, and total correctness is 80,0%. Conclusion. Balneotherapy causes multivariate changes in Popopvych’s step test, which accompanied by characteristic changes in certain parameters of autonomous nervous and immune systems as well as others functional and metabolic tests. Key words: Step test, HRV, Immunity, Metabolism, Balneotherapy, Children. INTRODUCTION Assessment of the functional state of the cardiovascular system, which limits the level of physical performance, is impossible without the use of load tests. The results of functional tests depend not only on the state of the heart and blood vessels, but also on the state of neuroregulatory mechanisms. Increasing the rhythm frequency during muscular work is mainly due to increased sympathetic effects on the heart and, to some extent, decreased parasympathetic [1,6,7]. The condition of the cardio-respiratory system is associated with the nervous, endocrine and immune systems, which collectively determine the level of adaptation of the body [9,11,16,17]. The purpose of this study is to analyze the effect on the step test of natural adaptogens and associated changes in a number of parameters of cardio-respiratory, autonomic nervous and immune systems and metabolism. MATERIAL AND METHODS The target was 80 children of both sexes, ages 10-17, arriving at the clinical sanatorium "Dzherelo" of Truskavets spa from radionuclide contaminated territories (density 137Cs 3793 kBq/m2). According to γ-spectrometry performed with a human radiation meter "Screensaver- 3M", the activity of incorporated 137Cs was 1,535 Bq/kg. The main subject of the study was the step test. As early as 1982, Dushanin SA et al [7] offered a modification of the step test to quantify the level of physical performance. The essence of the test is to rhythmically climb the stairs (height 37 cm) and lower it 90 times within 3 minutes. Both the pulse response to the load (whose power is 2,412 W/kg) and the speed of recovery are taken into account. The authors propose to calculate the index of physical performance (IPP) by the formula: IPP=(P1+P2+P3-200)/10, where P1 is pulse (heart rate) for 1 min in position sitting after 5 min of rest; P2 is heart rate for the first 10 sec after loading, multiplied by 6; P3 is heart rate for the first 10 sec of the second minute of recovery, multiplied by 6. 76 However, this formula does not take into account the actual time (t) spent by 90 ascents, so Popovych IL [28] has proposed a variant of the step test index (PSTI): PSTI=180•100/(HR1+HR2+HR3–200)•t State of autonomous nervous system was assessed by the method of HRV [2] using the “Cardio” device (Kyiv). However, on the basis of indicators of heart rate sitting and standing calculated Teslenko’s index [6]. In the same vein, an Anfimov’s corrective test was conducted to assess mental performance, as well as the classic Stange’s and Gench’s breath holding tests for inspiration and expiration, respectively. The level of hemoglobin was also determined. In the vein of functional-metabolic continuum concept [9] an oral glucose tolerance test (OGTT) was also performed [8]. Lymphocyte phenotypes (markers CD3, CD4, CD8, CD16, CD19) were determined by the method of indirect immunofluorescence reaction of binding of monoclonal antibodies of IHC "Sorbent" with visualization under a fluorescence microscope. Theophylline-resistant and theophylline-sensitive T-lymphocyte subpopulations, serum levels of G, A, M immunoglobulins (radial immunodiffusion method) and circulating immune complexes (polyethylene glycol precipitation method) were also determined. The state of the phagocytic link of immunity was judged by the activity of phagocytosis (phagocytic index), its intensity (microbial number) and completeness (killing index) against Staphylococcus aureus, with the calculation of bactericidal ability of neutrophils. Serum lysozyme activity was evaluated in the bacteriolysis test of Micrococcus lysodeikticus. To assess local immunity, lysozyme activity in mixed saliva was determined. Used the methodologies in the manuals [14,18,19]. Patients received a standard balneotherapeutic complex, the basis of which was drinking 3 ml/kg Naftussya water three times daily; mineral baths (concentration of Cl-SO4-Na-Mg salts 20- 30 g/L, t 36-37oC, duration 8-10 min) every other day, 8 procedures; ozokerite application to the lesion site (t 45oC, duration 20-30 min) every other day, 8 procedures; dietary nutrition; therapeutic exercise. Adaptogenic properties of the complex and its components have been proven by previous studies of the Truskavetsian Scientific School of Balneology [10- 13,16,17,20-27,29]. Results processed by methods of cluster and discriminant analyses, using the software package "Statistica 5.5". RESULTS AND DISCUSSION Preliminary analysis revealed multidirectional changes of the step-test, just as they are multidirectionally changed under the influence of adaptogens, in particular balneotherapy factors, parameters of various functional systems of the human body and animals [3,4,11,16,17]. For further analysis, the contingent surveyed was retrospectively divided into three qualitatively distinct groups. The largest group (53,7%) was the group of children with initially reduced or normal levels of PSI, in which it increased (Fig. 1), which is quite expected in line with the beneficial effect of balneotherapy on the body [16]. However, in 28,8% of children PSI remained stable at different initial levels, and in 17,5% even decreased, without falling below the lower limit of normal. In the adult contingent of the Truskavets spa it was shown earlier [28] that due to complex balneotherapy, the index of tachycardia-hypertensive reaction to 1,5 W/kg cycling load in 47,6% of patients increased by 16,6±2,4%, in 26,2% of persons they did not go beyond ±5%, instead of the remaining 26,2% the value of this index decreased by an average of 10,0±1,0%. In an experiment on female rats [28], it was found that swimming time to fatigue in 6 77 animals after 3 weeks of water Naftussya use increased from 13,0±1.4 min to 52,3±5,9 min, in 8 animals from 24,5±3,9 min to 37,3±5,9 min, instead of 4 rats with initial very high performance, adverse changes were observed, ie a reduction in swimming time from 61±7 min to 39±3 min. Thus, in the experimental model, a significant favorable actotropic effect was found in 33,3% of individuals, moderately favorable in 44,4% and unfavorable in 22,2% of cases. At the same time, the causation of actotropic effects by Naftussya water has been proved. Fig. 1. Individual levels of Popovych’s step-test index (PSI) before (X-axis) and after (Y-axis) course of adaptogenic balneotherapy. Green lines indicate normal limits Discriminant analysis [15] was subsequently applied to identify constellations of parameters that change are characteristic of each cluster. The program (forward stepwise) selected as characteristic changes in 2 parameters of autonomous nervous system, 2 functional, 4 metabolic and 3 immune parameters as well as blood trombocytes level. Of particular interest are a number of parameters that are not spent in the model (Tables 1 and 2). 78 Table 1. Discriminant Function Analysis Summary for Accompaniments of Changes in Popovych’s Step Test Step 15, N of vars in model: 15; Grouping: 3 grps Wilks' Lambda: 0,303; approx. F(30,1)=3,4; p<10-6 Changes in Variables currently in the model Changes in Popovych’s Step Test Parameters of Wilks’ Statistics PST- -0,10±0,02 (14) PST0 0,00±0,00 (23) PST+ +0,13±0,01 (43) Wilks Λ Par- tial Λ F- remove p- le- vel Tole ran- cy Leukocytes total, G/L -0,12±0,25 -0,29±0,17 +0,36±0,07 ,314 ,966 1,1 ,337 ,800 Stange’s Test, sec -2,6±3,6 -2,3±2,5 +5,9±1,8 ,350 ,867 4,9 ,011 ,799 Teslenko’s Test, points 0,2±0,6 -0,4±0,3 +1,0±0,3 ,410 ,739 11 ,000 ,600 Trombocytes, G/L -4±5 +11±6 +20±5 ,320 ,946 1,8 ,173 ,856 Glucosa a 1h OGTT, mM/L +0,15±0,31 +0,38±0,16 +0,63±0,10 ,314 ,964 1,2 ,315 ,702 Glucosa basal, mM/L 0,00±0,08 +0,08±0,09 +0,20±0,07 ,321 ,943 1,9 ,158 ,192 CIC, units +18±11 +14±6 -6±6 ,356 ,851 5,5 ,006 ,481 AMo HRV, % +2,5±2,5 0,0±2,1 -1,6±1,3 ,327 ,926 2,5 ,090 ,696 Pan Lymphocytes, % +4,5±2,2 -1,3±1,4 -0,9±1,3 ,325 ,933 2,3 ,113 ,678 Glycemia aft 2h OGTT, % +7,4±4,4 -1,6±4,7 -1,7±3,5 ,314 ,966 1,1 ,333 ,077 Phagocytose Index, % +18±3 +8±2 +10±1 ,358 ,847 5,7 ,005 ,782 Glucosa a 2h OGTT, mM/L +0,35±0,14 -0,05±0,17 +0,15±0,12 ,328 ,923 2,6 ,081 ,095 IgA Serum, g/L +0,15±0,28 -0,15±0,18 +0,24±0,11 ,316 ,958 1,4 ,262 ,772 Theoph Sensit T Lymph., % -3,6±1,2 +1,6±1,2 +1,0±1,1 ,336 ,903 3,4 ,041 ,514 MxDMn HRV, sec 0,00±0,05 +0,04±0,02 -0,01±0,02 ,354 ,856 5,3 ,007 ,619 Changes in Variables currently not in the model PST- -0,10±0,02 (14) PST0 0,00±0,00 (23) PST+ +0,13±0,01 (43) Wilks Λ Par- tial Λ F to en-ter p- le- vel Tole ran- cy CD8+ T Lymphocytes, % -1,8±0,8 +1,7±0,9 +1,2±0,8 ,297 ,980 ,63 ,536 ,051 CD16+ NK Lymphocytes, % -2,2±2,2 +2,6±1,0 +2,3±1,0 ,302 ,997 ,09 ,915 ,319 Theophyl Res T Lymph., % +7,5±2,9 +3,3±1,8 +3,4±1,8 ,297 ,979 ,65 ,525 ,668 CD4+ T Lymphocytes, % +4,3±1,5 +1,3±0,9 +1,4±0,9 ,297 ,979 ,65 ,525 ,561 Killing Index Neutroph, % +14±2 +5±2 +7±2 ,296 ,976 ,78 ,464 ,156 Bactericidity Neutr, 109 B/L +7,43±1,47 +4,31±0,77 +4,87±0,70 ,301 ,993 ,21 ,807 ,328 Hemoglobin, g/L +0,45±0,57 -0,11±0,34 +0,84±0,26 ,302 ,995 ,14 ,867 ,728 Erythroc Sedim Rate, mm/h 0,0±0,5 -1,6±0,4 -0,2±0,3 ,299 ,985 ,46 ,632 ,850 Anfimov’s Test, error/2 min -1,2±0,9 -2,3±0,5 -0,9±0,3 ,297 ,981 ,62 ,543 ,732 Hench’s Test, sec +1,6±4,4 -0,1±2,8 +5,3±1,3 ,295 ,974 ,83 ,441 ,596 Segmented Neutrophils, % -0,9±2,5 +3,4±1,5 -0,9±1,3 ,298 ,983 ,55 ,582 ,824 CD19+ B Lymphocytes, % +0,4±0,8 +1,6±0,6 +0,6±0,4 ,299 ,985 ,47 ,625 ,663 79 Table 2. Summary of Stepwise Analysis for Accompaniments of Changes in Popovych’s Step Test. The variables are ranked by criterion Lambda Changes in Variables currently in the model F to enter p- level Λ F- value p- level Leukocytes total, G/L 6,1 ,004 ,864 6,1 ,004 Phagocytose Index, % 4,9 ,010 ,765 5,4 10-3 Stange’s Test, sec 3,5 ,037 ,701 4,9 10-3 Teslenko’s Test, points 4,9 ,010 ,619 5,0 10-4 Glucosa basal, mM/L 3,1 ,050 ,570 4,7 10-5 CIC, units 2,6 ,084 ,532 4,5 10-5 Theophilline Sensitive T Lymph., % 3,0 ,057 ,491 4,3 10-5 MxDMn HRV, sec 3,3 ,042 ,448 4,3 10-6 Glucosa after 2h OGTT, mM/L 2,9 ,064 ,414 4,3 10-6 AMo HRV, % 2,2 ,122 ,389 4,1 10-6 Pan Lymphocytes, % 3,3 ,044 ,354 4,1 10-6 Trombocytes, G/L 1,5 ,240 ,339 3,9 10-6 IgA Serum, g/L 1,6 ,218 ,324 3,8 10-6 Glucosa after 1h OGTT, mM/L 1,0 ,365 ,314 3,6 10-6 Glycemia after 2h OGTT, % 1,1 ,333 ,303 3,4 10-6 Next, the 15-dimensional space of discriminant variables transforms into 2-dimensional space of canonical roots. The canonical correlation coefficient is for Root 1 0,738 (Wilks' Λ=0,303; χ2(30)=84; p<10-6) and for Root 2 0,578 (Wilks' Λ=0,666; χ2(14)=28; p=0,012). The major root contains 70,5% of discriminative opportunities and the minor is 29,5%. Table 3 presents standardized (normalized) and raw (actual) coefficients for discriminant variables. The calculation of the discriminant root values for each person as the sum of the products of raw coefficients to the individual values of discriminant variables together with the constant enables the visualization of each patient in the information space of the roots (Fig. 2). 80 Table 3. Standardized and Raw Coefficients and Constants for Canonical Variables Coefficients Standardized Raw Changes in Variables currently in the model Root 1 Root 2 Root 1 Root 2 Leukocytes total, G/L ,206 ,240 ,335 ,391 Phagocytose Index, % -,335 ,633 -,035 ,066 Stange’s Test, sec ,551 ,070 ,045 ,006 Teslenko’s Test, points ,881 ,186 ,448 ,095 Glucosa basal, mM/L ,518 -,669 1,215 -1,569 CIC, units -,717 -,295 -,020 -,008 Theophilline Sensitive T Lymph., % -,459 -,468 -,074 -,075 MxDMn HRV, sec -,497 -,541 -3,572 -3,888 Glucosa after 2h OGTT, mM/L ,103 1,551 ,133 2,001 AMo HRV, % -,429 ,128 -,047 ,014 Pan Lymphocytes, % -,344 ,320 -,043 ,040 Trombocytes, G/L ,285 -,239 ,009 -,008 IgA Serum, g/L ,306 -,095 ,372 -,115 Glucosa after 1h OGTT, mM/L -,092 -,374 -,116 -,473 Glycemia after 2h OGTT, % ,001 -1,156 ,000 -,053 Constants -,136 -,453 Eigenvalues 1,198 ,501 Cum. Properties ,705 1,000 Table 4 shows the correlation coefficients of PST changes (discriminant variables) with canonical discriminant roots and the cluster centroids of both roots. Table 4. Correlations Variables-Canonical Roots and Roots Means of changes in Variables for Clusters Changes in Variables currently in the model Correlations Variables-Roots Root 1 (70,5%) R 1 R 2 Leukocytes total, G/L ,357 ,105 Stange’s Test, sec ,316 ,002 Teslenko’s Test, points ,277 ,146 Trombocytes, G/L ,218 -,221 Glucosa after1h OGTT, mM/L ,199 -,134 Glucosa basal, mM/L ,163 -,079 CIC, units -,262 ,042 AMo HRV, % -,134 ,129 Pan Lymphocytes, % -,108 ,339 Glycemia after 2h OGTT, % -,077 ,193 Root 2 (29,5%) R 1 R 2 Phagocytose Index, % -,110 ,480 Glucosa after 2h OGTT, mM/L ,020 ,245 IgA Serum, g/L ,154 ,175 Theophiline Sensitive T Lymph., % ,093 -,336 MxDMn HRV, sec -,109 -,142 The above allows us to comment on Fig. 2 as follows. The localization of points representing children with PST growth in the right area of the axis of the first root reflects the 81 fact that it is accompanied by an increase in variables that correlate positively with the radical (Table 4), namely (Table 1): respiration delay time, improvement orthostatic test, increased blood levels of common leukocytes and platelets, basal glycemia, and one hour after oral glucose loading. On the other hand, the increase in PST is accompanied by a decrease in sympathetic tone and blood content of circulating immune complexes and a tendency to decrease the late hyperglycemic response to the loading of glucose and the proportion of pan-lymphocytes associated with the root inverse. We consider the described changes physiologically favorable. PST+- PST- PST+ Root 1 (71%) R oot 2 (29% ) -3 -2 -1 0 1 2 3 -4 -3 -2 -1 0 1 2 3 Fig. 2. Scatterplot of individual values of the first and second roots in which condensed information about of the changes in parameters of the members of the three clusters On the opposite side are localized projections on the axis of points, which represent children with stable PST and its negative dynamics, which are mixed, reflecting approximately the same decrease/increase in the above variables. These two groups of children are separated along the axis of the second radical. The upper position of points of children with negative PST changes reflects more pronounced, than in the neutral group, an increase in phagocytosis activity and a positive, as opposed to negative, dynamics of IgA levels and a late hyperglycemic reaction to glucose loading, on the one hand, and a decrease in the proportion of theophylline sensitive T-Lymphocytes and no increase in vagus tone, while in the other group both variables increase - on the other hand. Taking into account variables that are not included in the model (Table 1), we see that the negative dynamics of PST is accompanied by a decrease in the proportion of blood T-killers and natural killers in combination with an increase in T-helpers and bactericidal neutrophils, ie physiological assessment is ambiguous. Despite the individual ectopies, on the plane of the two discriminative roots, the members of the three groups are delimited statistically, which is documented by the calculation of Mahalanobis distances (Table 5). 82 Table 5. Squared Mahalanobis Distances between Clusters, F-values and p levels Clusters PST0 PST- PST+ PST0 0,00 4,61 5,40 PST- 2,06 0,025 0,00 6,72 PST+ 4,26 <10-4 3,64 <10-3 0,00 The same discriminant parameters can be used to identify the belonging of one or another person to one or another cluster. This purpose of discriminant analysis is realized with the help of classifying functions (Table 6). Table 6. Coefficients and Constants for Classification Functions of Clusters Changes in Variables currently in the model PST0 PST- PST+ p=,288 p=,175 p=,537 Leukocytes total, G/L -,737 ,061 ,298 Phagocytose Index, % ,116 ,257 ,096 Stange’s Test, sec -,047 -,038 ,053 Teslenko’s Test, points -,547 -,381 ,484 Glucosa basal, mM/L ,252 -3,141 1,535 CIC, units ,034 ,018 -,015 Theophilline Sensitive T Lymph., % ,158 ,005 -,062 MxDMn HRV, sec 7,241 -,682 -3,569 Glucosa after 2h OGTT, mM/L -1,479 2,724 ,460 AMo HRV, % ,053 ,086 -,036 Pan Lymphocytes, % ,038 ,126 -,020 Trombocytes, G/L ,012 -,005 ,025 IgA Serum, g/L -,343 -,613 ,352 Glucosa after 1h OGTT, mM/L 1,487 ,499 ,849 Glycemia after 2h OGTT, % ,034 -,076 -,009 Constants -2,813 -4,842 -2,378 The correct classification of the positive actotropic effect is 93,0%; neutral 65,2%; negative 64,3%, and total correctness is 80,0% (Table 7). Table 7. Classification Matrix for Clusters Rows: Observed classifications; Columns: Predicted classifications Percent PST+ PST0 PST- Clusters Correct p=,538 p=,287 p=,175 PST+ 93,0 40 2 1 PST0 65,2 5 15 3 PST- 64,3 2 3 9 Total 80,0 47 20 13 To compare accuracy, we recall that in adult patients were selected for inclusion in model 10 variables from 72 counts (in order of decreasing F-value): tachycardia-hypertensive load index, Klimov atherogenic factor, cholesterol content VLD LP, sodium content erythrocytes, activity of Na, K-ATPase erythrocyte shadows, level of diastolic and systolic blood pressure; the plasma potassium and urates content, as well as the Ruzhylo-Popovych’s contractile activity index of 83 myocardium. The correct classification of the positive actotropic effect is 81,0%; neutral 60,0%; negative 72,7%, and total correctness is 75,6% [28]. In the next article, the possibility of predicting the diversity of actotropic effects of adaptogenic balneotherapy will be considered. 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Actotropic Effects of Balneotherapeutic Complex of Truskavets spa [in Ukrainian]. Кyiv. Computerpress; 2003: 131 p. 29. Sydoruk NO, Zukow W. Differences between the effects of water Naftussya from fields of Truskavets’ and Pomyarky on the parameters of the EEG, HRV, immunity and metabolism. Journal of Education, Health and Sport. 2019; 9(1): 287-293. Table 3 presents standardized (normalized) and raw