Wojciech Stojek

Active dynamic thermography in cardiosurgery

The aim of this study is development and analysis of applicability of new tools for continuous inspection of open-heart cardio-surgical interventions, suitable in clinical environment. Our notice is concentrated on IR-thermal imaging for monitoring of invasive surgical interventions as CABG in treatment of ischaemia and other heart malfunctions. In this presentation preliminary results of in-vivo experiments on animals with the use of IR thermal camera and newly developed procedures of active dynamic thermography are discussed.

Small doses of morphine can enhance NK cell cytotoxicity in pigs.

The effect of small and moderate doses of morphine (MF) on NK cell lytic activity (cytotoxicity, NKCC) ((51)Cr release test) and the number of circulating large granular lymphocytes (LGL) was evaluated in i.v. catheterized Pietrain crossbred pigs. Simultaneously, plasma cortisol (COR) (RIA method) was measured. Blood samples were collected 15, 60, 120, 180, and 240 min after i.v. injections of 0.5, 1.0 and 5.0 mg/kg of MF alone or MF pretreated with naloxone (NX, 1.0 mg/kg, i.v., 15 min before MF). It was found that MF induced dose- and time-dependent changes of NKCC. MF in a dose of 0.5 mg/kg evoked 4-fold increase in NKCC (in comparison to saline) without changes in the number of LGL/NK cells. Higher MF doses (1.0, 5.0 mg/kg) induced an early increase (up to 300Delta% and 29Delta%, respectively) followed by a decrease in cytotoxicity (to -76Delta% after 5.0 mg/kg), and in LGL number (-36Delta% after 5.0 mg/kg). These effects were concomitant with a marked rise in plasma COR (up to 234Delta% after 0.5 mg/kg and 567Delta% after 5.0 mg/kg of MF). NX pretreatment blocked all the changes in cytotoxicity but not in the LGL cell number and COR concentrations. The results indicate that MF, besides having well known immunosuppressive effects, can also enhance NKCC through the opioid receptors-dependent manner. The enhancement of cytotoxicity appears as a purely functional change independent of the recirculation of NK cells which occurs despite the high plasma concentrations of COR.

Restraint effects on stress-related hormones and blood natural killer cell cytotoxicity in pigs with a mutated ryanodine receptor

A mutation in the ryanodine receptor gene (RYR1) of the calcium release channel is responsible for increased stress susceptibility in pigs. In the present study, the relation of a mutation in RYR1 with the neuroendocrine (stress-related hormone) response and the immune defense represented by natural killer cell cytotoxicity (NKCC) during a 4-h restraint and recovery phase in 60 male pigs was investigated. Blood samples were collected from pigs previously divided into RYR1 genotypes (nn, Nn, NN), based on PCR amplification and restriction analyses. The blood samples collected during the restraint and recovery phases of the experiment were used to determine NKCC ((51)Cr-release assay), large granular lymphocyte number (hematologic method), and plasma concentrations of prolactin (PRL), GH, ACTH, and cortisol (COR) (by specific RIA). The greatest degree of NKCC response (P < 0.05) to restraint stress relative to controls was observed for the stress-susceptible homozygote group (nn). Measures of stress-related hormones were positively correlated with NKCC during the entire experimental period (P < 0.001 for all investigated hormones) in the nn group. Immunostimulatory effects in the early (0-60 min) phase of restraint were associated with increased hormone responses, especially PRL and GH. In the late (180-240 min) phase of stress and the recovery phase (480 min), a decrease in immune response was accompanied by an elevated COR response in all RYR1 genotypes. Moreover, divergent responses of both PRL (greatest in nn, P < 0.001) and GH (greatest in NN, P < 0.001) to the 4-h restraint were observed. Our results suggest that stress-susceptible RYR1-mutated homozygotes develop a greater level of immune defense, including cytotoxic activity of NK cells, and accompanied by more pronounced stress-induced changes in neuroendocrine response than stress-resistant heterozygous (Nn) and homozygous (NN) pigs.

Investigation of skin burns basing on active thermography

Use of dynamic thermography for assessment of burns is discussed. Animal in-vivo experiments are presented. Sets of burns were inflicted on the backs of eight domestic pigs. Thermographic measurements of burns with different depths of affected tissue (from first to third-degree burns) were correlated with histopathologic analysis of lesions. The results show that dynamic thermography might be applied as a simple, noninvasive and nonstressful tool for patient diagnosis. Further analysis of dynamic pictures gives the first estimate of the depth of a lesion.

Natural killer cell cytotoxicity, cytokine and neuroendocrine responses to opioid receptor blockade during prolonged restraint in pigs

This study evaluated porcine natural killer cell cytotoxicity (NKCC), plasma cytokines including interleukin (IL) 1β, IL-6, IL-10, IL-12 and tumor necrosis factor-α and plasma stress-related hormones including prolactin (PRL), growth hormone (GH), β-endorphin (BEND), ACTH and cortisol (COR) during a 4h restraint and recovery phase after saline or naloxone (1mg/kg BW) administration. The restraint preceded with saline altered NKCC and IL-12 concentration (an early from 15 to 60 min increase followed by a decrease) and increased other measured cytokines and hormones concentrations. Naloxone pretreatment blocked the suppressive effects of the restraint on NKCC and IL-12 and altered IL-10, IL-6, TNF-α, PRL and ACTH concentrations. Furthermore, in naloxone-injected pigs, a positive correlation was found between NKCC and all measured cytokines (with the exception of IL-6) and BEND, ACTH and COR. Results suggest that naloxone-sensitive opioid pathways could influence the mechanisms underlying the immune system (including NKCC) response during stress.

Thermographic and electrical measurements for cardiac surgery inspection

The aim of this paper is to analyze the validity of new methods applicable for cardiac surgery intraoperation inspection. In vivo measurements on pigs using passive and active thermography as well as electroimpedance spectroscopy applied to investigation of the heart tissue properties during open chest cardiac surgery are related. The measurement results clearly show that each of proposed modalities is giving different information valuable for evaluation of the heart muscle biological properties. This information may have very high importance in terms of intra-operation inspection as well as for decisions of proper medical interventions. A series of cardiac operations on anaesthetized pigs involving a controlled heart infarct followed by extensive histopathologic study allowed the authors to make an objective evaluation of the value of the discussed methods.

The effects of ryanodine receptor 1 (RYR1) mutation on plasma cytokines and catecholamines during prolonged restraint in pigs.

In the current study, plasma cytokines, including interleukin (IL) 1, IL-2, IL-6, IL-10, IL-12, interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) and catecholamines (adrenaline and noradrenaline) were evaluated during 4h restraint and recovery phase in 60 male pigs. Blood samples were collected from three groups of pigs representing different RYR1 genotypes, namely NN homozygotes (wild-type), Nn heterozygous and nn homozygous (mutant). The 4h restraint evoked an increase in plasma cytokine concentrations in each of the RYR1 genotype groups. During the restraint, the greatest concentrations of plasma IL-6, IL-10, IL-12 and TNF-α in nn homozygous pigs and IFN-γ in NN homozygous were observed. Interleukin 1, IL-2, IL-10, and TNF-α measures were positively intercorrelated in each of the three RYR1 genotype group. A positive correlation was seen between all measured cytokines (with the exception of IL-6) and plasma catecholamine concentrations in Nn heterozygous and nn homozygous pigs. The results suggest that of the cytokine parameters evaluated, IL-6, IL-10, IL-12 and TNF-α of the nn homozygous group seem to show a stronger stress-related response as compared with those of the other two (NN and Nn) groups.

Thermal monitoring of the myocardium under blood arrest preliminary study

A method for monitoring the state of the myocardium during cardiosurgical interventions based on thermal IR imaging is presented. This method called active dynamic thermography (ADT) uses an external excitation source to induce thermal transient processes in a tested object. Recording time series of thermograms allows to calculate parametric images - distribution of the thermal time constant at the visible surface of the myocardium - correlated to the physiology state of tested tissues. In the experiment performed the blood arrest in the left ventricle (causing in longer period heart infarct) was generated by clamping the left anterior descending artery (LAD). During LAD occlusion the ADT procedure was applied using cooling external excitation source. In summary, the calculated time constant images provide data of the tested structure and functional information of myocardium infarct. This allows to track changes in blood flow in the myocardium and inspection of the quality of the intervention during cardiosurgical procedures.

The effects of ryanodine receptor (RYR1) mutation on natural killer cell cytotoxicity, plasma cytokines and stress hormones during acute intermittent exercise in pigs

Stress susceptibility has been mapped to a single recessive gene, the ryanodine receptor 1 (RYR1) gene or halothane (Hal) gene. Homozygous (Hal(nn)), mutated pigs are sensitive to halothane and susceptible to Porcine Stress Syndrome (PSS). Previous studies have shown that stress-susceptible RYR1 gene mutated homozygotes in response to restraint stress showed an increase in natural killer cell cytotoxicity (NKCC) accompanied by more pronounced stress-related hormone and anti-inflammatory cytokine changes. In order to determine the relationship of a RYR1 gene mutation with NKCC, plasma cytokines and stress-related hormones following a different stress model - exercise - 36 male pigs (representing different genotypes according to RYR1 gene mutation: NN, homozygous dominant; Nn, heterozygous; nn, homozygous recessive) were submitted to an intermittent treadmill walking. During the entire experiment the greatest level of NKCC and the greatest concentrations of interleukin (IL-) 6, IL-10, IL-12, interferon (IFN-)γ and tumor necrosis factor-α and stress-related hormones (adrenaline, prolactin, beta-endorphin) were observed in nn pigs, and the greatest concentration of IL-1 and growth hormone in NN pigs. Immunostimulatory effects of intermittent exercise on NKCC in nn pigs were concomitant with increases in IL-2, IL-12 and IFN-γ, the potent NKCC activators. Our findings suggest that stress-susceptible pigs RYR1 gene mutated pigs develop a greater level of NKCC and cytokine production in response to exercise stress. These results suggest that the heterogeneity of immunological and neuroendocrine response to exercise stress in pigs could be influenced by RYR1 gene mutation.

IR-thermal monitoring of cardiosurgery interventions

Results of a research project devoted to study of quality of open heart surgical interventions are presented. The aim of the project was analysis of cardiosurgery therapeutic procedures in terms of reduction of inter- and post- operative complications. The objective of the research was development of quantitative diagnostic and measurement methods for evaluation of open-heart cardiosurgery interventions but also other applications. Prototype instrumentation and software for application in clinical environment and new measurement and visualization procedures for presentation of results were developed. The results of this project are here presented. Diagnostic thermal parameters were analysed from the point of view of practical importance in evaluation of cardiosurgery interventions. The applied methodology is evaluated using in vivo animal experiments. High importance of thermal imaging for evaluation of open-heart cardiosurgery interventions is underlined. Some clinical results are also presented.