Novi dokazi ukazuju da je antinociceptivno djelovanje botulinum toksina tipa A (BT-A) središnjeg porijekla. U ovom doktorskom radu smo provjerili ovu pretpostavku u ranije nedovoljno istraženim ...oblicima boli, istraživali interakciju sa središnjim neurotransmitorima kao mogući mehanizam djelovanja te pokušali pobliže utvrditi mjesto djelovanja toksina u središnjem živčanom sustavu (SŽS).
Ispitivanja su izvedena na mužjacima Wistar štakora. U upalnoj, neuropatskoj i bilateralnoj mišićnoj boli ispitan je učinak selektivnih i neselektivnih antagonista opioidnih i GABAA receptora, primijenjenih sistemski, spinalno ili supraspinalno, na antinociceptivno djelovanje periferno (supkutano u šapu) primijenjenog BT-A. U tkivu kralješnične moždine ispitana je aktivacija neuronalnih i glija stanica, ekspresija mRNA proupalnih citokina i μ-opioidnih receptora te ekspresija Leu/Met-enkefalina metodama imunofluorescencije i lančane reakcije polimerazom s reverznom transkripcijom. Istraživano je antinociceptivno djelovanje BT-A nakon periferne, spinalne i supraspinalne primjene te je imunofluorescencijom ispitana enzimska aktivnost BT-A u tkivu SŽS-a.
Opioidni i GABAA antagonisti su ovisno o dozi, sistemski i intratekalno, ali ne i supraspinalno, poništili antinociceptivno djelovanje BT-A u svim ispitanim modelima. Učinak antagonista bio je kratkotrajan. BT-A je smanjio neuronalnu aktivaciju u dorzalnom rogu kralješnične moždine, što su antagonisti blokirali. BT-A je smanjio bol u dosad neistraženim modelima visceralne boli (peritonitis, kolitis). Bilateralno antinociceptivno djelovanje BT-A posljedica je prisutnosti toksina samo na ipsilateralnoj strani. BT-A je smanjio bol nakon periferne i intratekalne primjene, dok primijenjen supraspinalno (cisterna magna, moždane komore) nije djelovao, unatoč nalazu njegove enzimske aktivnosti u pojedinim regijama mozga uključenima u nocicepciju.
BT-A ima segmentalno antinociceptivno djelovanje spinalnoj razini, uz neizravnu aktivaciju endogenog opioidnog i GABA-ergičkog sustava. Ovi bi nalazi mogli biti važni za klinička ispitivanja potencijalno korisnih sinergističkih interakcija s konvencionalnim analgeticima i drugim lijekovima te usmjeriti klinička ispitivanja na nove indikacije i nove načine primjene, poput intratekalne.
Novel evidence suggests that the antinociceptive effect of botulinum toxin type A (BT-A) is of central origin. In this doctoral thesis, we verified this assumption in previously insufficiently investigated types of pain, investigated the interaction with central neurotransmitters as the possible mechanism of action and tried to determine the site of the toxin’s action within the central nervous system (CNS).
Male Wistar rats were used in experiments. We examined the effect of selective and nonselective opioid and GABAA antagonists, applied systemically, spinally and supraspinally, on antinociceptive effect of peripherally (subcutaneously into hind paw) applied BT-A in inflammatory, neuropathic, and bilateral pain. Neuronal and glial cells’ activation, proinflammatory cytokines’ and μ-opioid receptors mRNA expression, and Leu/Met-enkephalin protein expression were analyzed using immunofluorescence and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) in lumbar spinal cord tissue. Further, we investigated the antinociceptive effect of BT-A following peripheral, spinal and supraspinal application. In parallel, enzymatic activity of BT-A in CNS tissue was examined using immunofluorescence.
Opioid and GABAA antagonists, applied systemically and intrathecally, dose-dependently abolished the antinociceptive effect of peripheral BT-A in all tested models, while no effect was observed following their supraspinal application. The effect of antagonists was short-lasting. BT-A reduced neuronal activation in dorsal horn, which was abolished by both, opioid and GABAA antagonist. BT-A diminished pain in, yet uninvestigated, models of visceral pain (peritonitis and colitis). BT-A’s bilateral antinociceptive action occurs after toxin’s presence on ipsilateral side only. BT-A reduced pain after peripheral and intrathecal application, but not after application in cisterna magna or cerebral ventricles, despite of its enzymatic activity in brain regions involved in nociception.
BT-A has segmental antinociceptive effect at the spinal level, which involves an indirect activation of endogenous opioid and GABA-ergic systems. These findings might guide clinical investigations of potentially useful additive or synergistic effects with conventional analgesics and direct clinical trials for novel indications and the routes of application, like intrathecal.
U fiziološkom odgovoru na stres i kontroli stresnog odgovora uglavnom sudjeluju dvije komponente neuroendokrinog sustava, simpatičko-medularna (SAM) os, koja regulira prvi odgovor na stres i os ...hipotalamus-adenohipofiza-kora nadbubrežne žlijezde (HPAC), čija se aktivacija uočava 15 do 20 minuta nakon stresnog podražaja. Cilj ovog istraživanja bilo je ispitati bazalnu, fiziološku aktivnost SAM i HPAC osi te njihovu povezanost s nizom kovarijabli (spol, tip škole, turnus pohađanja nastave, ritam spavanja i budnosti te demografska, akademska i društvena obilježja) u zdravih učenika koji pohađaju završne razrede srednjih škola. Specifični ciljevi uključivali su ispitivanje koncentracije kortizola i aktivnosti alfa-amilaze kao salivarnih biljega odgovora na stres te istraživanje njihove povezanosti sa subjektivnim procjenama razine stresa i sa strategijama suočavanja sa stresom, kao i s polimorfizmima jednog nukleotida (SNP) gena za mineralokortikoidni i glukokortikoidni receptor (rs5522, rs6189, rs6190) te za vezni protein 5 za FK506 (rs1360780). U istraživanje su bila uključena 903 maturanta gimnazija i strukovnih škola iz četiri najveća grada u Hrvatskoj (Zagreb, Split, Rijeka i Osijek). Uzorci sline prikupljeni kod kuće, tijekom jednog radnog dana u tri vremenske točke (po buđenju, 30 do 45 minuta nakon buđenja i neposredno prije lijeganja), uzeti su za analizu koncentracija kortizola dok se aktivnost alfa-amilaze mjerila samo u jednom uzorku, neposredno po buđenju. Kako bi se ispitao cirkadijalni ritam lučenja kortizola analizirani su i indeksi lučenja kortizola (CAR, DCD, AUCG). Učenice su imale statistički značajno veće koncentracije kortizola u jutarnjim satima, dok se koncentracije kortizola neposredno prije lijeganja nisu razlikovale među spolovima. Učenice imaju različite obrasce priprema za izazove koji ih očekuju tijekom nadolazećeg dana u odnosu na učenike, imaju značajno viši CAR, izraženiji DCD, veći AUCG u odnosu na učenike koji su imali statistički značajno veću aktivnost alfa-amilaze. Turnus pohađanja nastave, koji svakako posljedično utječe na ritam spavanja i trajanje budnosti, pokazao se kao prediktor lučenja kortizola. Statistički značajne razlike većine varijabli kortizola potvrđene su između učenika koji pohađaju različite turnuse nastave (SCC30-45, CAR, DCD i AUCG). Učenici koji su se budili ranije i imali duže trajanje budnosti imali su veći CAR, niži DCD i veći AUCG. Nisu potvrđene statistički značajne razlike niti jedne ispitivane varijable u učenika koji pohađaju različite tipove škola (gimnazije vs. strukovne škole). Također, statistički značajne razlike između učenika i učenica bile su uočene i u subjektivnoj procjeni stresa, suočavanju te njihovoj povezanosti s koncentracijama i indeksima kortizola. Zabrinutost za budućnost su oba spola procijenila kao najstresniju domenu života, dok su učenice imale statistički značajno veću razinu ukupnog stresa u odnosu na učenike te su više koristile aktivno suočavanje sa stresom. U ovom su istraživanju pronađene statistički značajne razlike povezanosti alela G polimorfizma rs 5522 s višom koncentracijom kortizola po buđenju (P=0,036), nižim CAR-om (P=0,021), dok je povezanost s DCD-om bila granična (P=0,050). Može se pretpostaviti kako učenici s dokazanim alelom G imaju niži CAR te zbog toga pokazuju lošiju pripremljenost za stresne izazove u nadolazećem danu, unatoč nešto izraženijim vrijednostima DCD-a. Ovo istraživanje ukazuje na potrebu definiranja protokola prikupljanja uzoraka sline u nekliničkim uvjetima i metodoloških smjernica za populacijska istraživanja, ponajprije zbog postojanja niza kovarijabli koje utječu na mjerenje koncentracije kortizola i aktivnosti alfa-amilaze. Dobiveni rezultati doprinose saznanjima o spolnim razlikama u funkciji HPAC osi, koje bi se trebale primijeniti i tijekom provedbe preventivnih mjera u adolescenata te ranijeg otkrivanja skupina koje su posebno osjetljive na stresne poticaje.
Physiological response to stress and the control of the stress response is mediated by two major components of the neuroendocrinological systems, sympathetic nervous (SAM) system and hypothalamic-pituitary-adrenal (HPA) axis. In the stress response, the SAM axis is first activated. About fifteen to twenty minutes following the SAM activation, the HPA axis is activated. The aim of the study was to examine basal, physiologically activity of the SAM and HPAC axis activity and to determine associations of various covariates (gender, school type, school shift, sleep-wake rhythm, demographic, academic and life style characteristics) with altered daily salivary cortisol profiles in healthy students attending finishing grades of secondary schools. Specific aims of the study were to analyse concentration of cortisol and activity of alpha- amylase, as salivary markers of stress reaction, to determine association of salivary markers of stress with stress perception and coping, and to determine associations of salivary markers of stress with single nucleotide polymorphisms (SNP) in the genes for the mineralocorticoid and glucocorticoid receptors (rs5522, rs6189, rs6190) and FK506 binding protein 5 (rs1360780). The study included 903 secondary school students enrolled in the finishing classes of gymnasiums and vocational schools from the four largest cities in Croatia (Zagreb, Split, Rijeka and Osijek). The saliva samples were collected at students’ homes, over the course of one weekday. Salivary cortisol was sampled at three time points: at awakening, 30 to 45 after wakening and at bedtime. Salivary alpha-amylase was sampled at awakening. In order to analyse the circadian rhythm of salivary cortisol secretion, three indexes were analysed (CAR, DCD and AUCG).Females had higher morning concentrations of salivary cortisol than males, whereas bedtime cortisol concentrations were not different in females and males. Females had different salivary cortisol profile and different mechanisms for preparing for the anticipated stress of the upcoming day, higher CAR, steeper DCD and larger AUCG than males. On the other side, males had higher activity of salivary alpha-amylase. School shift, and therefore the sleep-wake up rhythm, was an important predictor of the secretion of salivary cortisol. There were statistically significant differences in two school shifts with respect to the majority of salivary cortisol measures (SCC30-45, CAR, DCD and AUCG). Students who woke-up earlier and were longer awake had larger CAR, flatter DCD and larger AUCG. Students from two types of schools (gymnasiums vs. vocational schools) did not differ in any of the measured salivary variables. Furthermore, female and male students differed significantly in stress perception, coping and associations between stress perception, coping and salivary cortisol concentrations and indexes. Future was rated as the most stressful problem domain in both sexes. The perception of total stress was higher in females than males, and females used more active coping than males. Finally, we found statistically significant differences in associations between G allele of rs 5522 polymorphism and higher cortisol concentration at awakening (P=0.036), lower CAR (P=0.021) and higher DCD (P=0.050), but at borderline. It can be hypothesized that the G allele is associated with the lower CAR and therefore, decreased ability of preparing for the anticipated stress (despite borderline association with the steeper DCD). This study points to the need for defining the protocol for salivary sampling in non-clinical conditions and developing methodological guidelines for population-based research, primarily due to the number of co-variables that affect the concentration of salivary cortisol and the activity of salivary alpha-amylase. Obtained results contribute to better understanding of sex differences in the functioning of the HPA axis, which should be considered in the prevention of various psychopathological conditions in adolescence and used for earlier identification of vulnerable groups at high risk of stress-related health problems.
Kronična opstrukcijska plućna bolest (KOPB) je složeni poremećaj koji zahvaća pluća, ali
i sistemski odjeljak. Naše se istraživanje temeljilo na hipotezi da je u bolesnika s KOPB-om
prisutna ...sistemska upala i sistemski oksidacijski stres. Sukladno tome, cilj ovog istraživanja
bio je istražiti sistemske pokazatelje oksidacijskih i upalnih promjena u bolesnika sa stabilnim
KOPB-om te ispitati povezanost sistemskih antioksidansa s poremećajem funkcije pluća i
biljezima sistemske upale. Ispitali smo i dijagnostičku učinkovitost biljega sistemskog
oksidacijskog stresa u razlikovanju zdravih i oboljelih od KOPB-a.
Sistemski biljezi oksidacijskog stresa (albumin, transferin, ceruloplazmin, ukupni bilirubin,
slobodne tiolne skupine, malondialdehid (MDA), paraoksonazna i arilesterazna aktivnost
paraoksonaze 1 (PON1), te ekspresija i aktivacija unutarstaničnih signalnih molekula Hsp27,
Hsp70, ERK, JNK i p38) određeni su u 106 bolesnika sa stabilnim KOPB-om i 45 zdravih
ispitanika. Ispitane su i njihove povezanosti s biljezima sistemske upale (CRP, fibrinogen,
ukupni leukociti), pušenjem i pokazateljima funkcije pluća (FEV1 i FEV1/FVC).
Bolesnici s KOPB-om su imali povišene koncentracije ceruloplazmina i MDA, snižene
koncentracije albumina, transferina i tiola, te snižene obje ispitane aktivnosti PON1.
Koncentracije ukupnog bilirubina nisu se razlikovale usporedbom ispitivanih skupina.
Ceruloplazmin je pokazao pozitivnu korelaciju s CRP-om i fibrinogenom. Albumin i
transferin su pokazali negativnu korelaciju s CRP-om, te pozitivnu korelaciju sa slobodnim
tiolima. Transferin je negativno korelirao s fibrinogenom. Jedini pokazatelj povezan s
funkcijom pluća bio je MDA. Usporedbom pušača, bivših pušača i nepušača iz skupine
bolesnika s KOPB-om nisu nađene razlike u koncentracijama promatranih biljega
oksidacijskog stresa i upale. Povišene koncentracije ceruloplazmina su najsnažniji prediktor
prisutnosti KOPB-a. Model koji uključuje ceruloplazmin, albumin, MDA i arilesteraznu
aktivnost PON1 te biljege sistemske upale pokazao je najbolju dijagnostičku učinkovitost u
predviđanju KOPB-a (AUC (95%CI) = 0,96 (0,92 – 0,99)). Predloženi bi model mogao
ispravno predvidjeti prisutnost KOPB-a u 89% bolesnika. Također, naši su rezultati ukazali da
je razina ekspresije Hsp27 i Hsp70 u leukocitima periferne bila je najniža u pušača s KOPBom.
Ekspresija svih ispitivanih MAPK (ERK, JNK i p38) u perifernim leukocitima bolesnika
s KOPB-om nije se razlikovala u usporedbi sa zdravim ispitanicima. Aktivacija ERK bila je
značajnija kod zdravih i bolesnih nepušača, dok je aktivacija JNK i p38 bila najizraženija u
pušača s KOPB-om.
Rezultati su ukazali da prisutnost bolesti i pušenje utječu na ispitane unutarstanične
signalne molekule. Bolje razumijevanje ovih molekularnih mehanizama moglo bi pomoći u
dijagnozi i pronalaženju novih terapijskih meta za KOPB. Dijagnostičke karakteristike
predloženog modela koji kombinira koncentracije biljega sistemske upale i sistemskog
oksidacijskog stresa ukazuju da bi se on mogao koristiti kao vrijedan alat u razlikovanju
zdravih ispitanika i bolesnika s KOPB-om.
Background: Chronic obstructive pulmonary disease (COPD) is a complex disorder
affecting the lungs and the systemic compartment. We hypothesized that systemic
inflammation and systemic oxidative stress are present in patients with COPD. Therefore, we
aimed to investigate markers of systemic oxidative and inflammatory alterations in patients
with stable COPD, and to test the association of systemic antioxidants with indicators of lung
function and systemic inflammation. The diagnostic accuracy of systemic oxidative stress
parameters in distinguishing between healthy subjects and patients with COPD was also
evaluated.
Materials and methods: Systemic oxidative stress markers (albumin, transferrin,
ceruloplasmin, total bilirubin, thiols, malondialdehyde (MDA), paraoxonase and arylesterase
activity of paraoxonase 1 (PON1), and the expression and activation of intracellular signalling
molecules Hsp27, Hsp70, ERK, JNK i p38) were assessed in 106 stable COPD patients and
45 healthy subjects. Their association with systemic inflammatory markers (CRP, fibrinogen,
total leukocytes), smoking status and lung function parameters (FEV1 i FEV1/FVC) was
investigated.
Results: Higher ceruloplasmin and MDA concentrations, and lower albumin, transferrin,
thiols and PON1 activities (paraoxonase and arylesterase) were found in patients with COPD.
Total bilirubin concentrations were similar in the studied groups. Ceruloplasmin showed a
positive correlation with CRP and fibrinogen. Albumin and transferrin showed a negative
correlation with CRP, and a positive corelation with thiols. Transferrin negatively correlated
with fibrinogen. Only MDA showed an association with pulmonary function. No differences
were found comparing concentrations of oxidative stress and inflammatory markers between
COPD patients subdivided according to their smoking status. Ceruloplasmin was the strongest
single predictor of COPD. The model combining ceruloplasmin, albumin, MDA, arylesterase
PON1 activity, and markers of systemic inflammation demonstrated very good diagnostic
performances (AUC (95%CI) = 0,96 (0,92 – 0,99)). The proposed model correctly identifies
89% of patients with COPD. In addition, our results showed that the decrease in expression of
peripheral blood leukocytes' Hsp27 and Hsp70 was the most prominent in COPD smokers.
Expression levels of all three MAPKs investigated was not altered in leukocytes of COPD
patients compared to healthy subjects. However, ERK activation was stimulated in healthy and COPD non-smokers, while JNK and p38 activation was the most pronounced in COPD
smokers.
Conclusions: Our results showed that COPD and smoking affect the intracellular
signalling pathways investigated. Improved understanding of these molecular mechanisms
could help identify novel targets for diagnosis and therapeutic interventions in COPD.
Diagnostic characteristics of the proposed model, obtained by combining markers of systemic
inflammation and systemic oxidative stress, suggest its potential value as an additional tool in
COPD diagnosis.
Kardiovaskularne bolesti uzrokovane aterosklerozom predstavljaju značajan uzrok smrtnosti u hemodijaliziranih
(HD) bolesnika. Bolesnici koji se liječe HD imaju i do 30 puta veću stopu smrtnosti ...uzrokovanu kardiovaskularnim
bolestima. Rizični čimbenici uključeni u patogenezu ateroskleroze u HD bolesnika pacijenata uključuju dijabetes,
hipertenziju, dislipidemiju, pušenje, oksidativni stres, upalu te endotelnu disfunkciju. Budući da ateroskleroza ima
dugačku subkliničku fazu veoma je važno da patološki procesi budu otkriveni čim ranije, dok je bolest još u
asimptomatskoj fazi. Povećana debljina intime-medije (IMT) prepoznata je kao rani indikator subkliničke
ateroskleroze, kako u općoj populaciji tako i u HD-ih bolesnika te je povezana s tradicionalnim i novijim
kardiovaskularnim čimbenicima rizika. Nekoliko je studija dokazalo da je karotidna ateroskleroza povezana s
adhezijskim molekulama te omentin-1 proteinom. Ciljevi ovog rada bili su sljedeći: a) istražiti povezanost sICAM-1,
sVCAM-1, omentin-1 proteina te ostalih netradicionalnih rizičnih čimbenika sa subkliničkom aterosklerozom; b)
ispitati dijagnostičku vrijednost ovih specifičnih markera u otkrivanju subkliničke ateroskleroze i c) ispitati njihovu
ulogu kao prediktora smrtnosti u asimptomatskih bolesnika.
Počevši od studenog 2011. godine, kohorta od 210 HD-ih bolesnika sudjelovala je u trogodišnjem istraživanju.
Ispitanici su bili podijeljeni u tri skupine ovisno o prisutnosti ateroskleroze. Ateroskleroza je dijagnosticirana na
temelju mjerenja debljine intima-medija karotidne arterije. Uzorkovanje krvi izvršeno je na početku studije te svakih
12 mjeseci iza toga sve do kraja istraživanja. Ehokardiografska mjerenja učinjena su na početku i na kraju studije.
Dokazana je slaba korelacije između IMT i sICAM-1 (r=0,39, P=0,001), sVCAM-1 (r=0,27, P=0,015) i omentin-1
proteina (r=-0,25, P=0,020). Omentin-1 protein je pokazao dobru korelaciju s parametrima sistoličke i dijastoličke
funkcije lijeve klijetke (r=0,52, P=0,001 i r=0,51, P=0,001). Multivarijantna analiza je pokazala kako su koncentracije
sICAM-1 i omentin-1 proteina snažni neovisni pretkazatelji IMT (P=0,031 i P=0,010). Coxova analiza
proporcionalnih rizika pokazala je da su koncentracija sICAM-1 i omentin-1 proteina snažni pretkazatelji
kardiovaskularne smrti (HR=1,85, CI=1,18-2,32, P=0,021 i HR=4,14, CI=1,38-12,1, P=0,004,) te da uzastopno
mjerenje koncentracija sICAM-1 i omentin-1 proteina predstavlja jake prediktore IMT progresije (HR=1,98, 95%
CI=1,21-2,38, P<0,002 i HR=2,91, 95% CI=1,57-4,72, P<0,001).
Naša studija je pokazala kako su koncentracije ICAM-1 I omentin-1 proteina jaki pretkazatelji kardiovaskularne
smrti u asimptomatskih HD bolesnika. Porast koncentracije ICAM-1 te pad koncentracije omentin-1 proteina imaju
ključnu ulogu u ranoj progresiji ateroskleroze.
Atherosclerotic cardiovascular complications represent significant cause of mortality in hemodialysis (HD)
patients. Patients undergoing HD have up to 30 times higher incidence of cardiovascular disease mortality. Risk
factors involved in pathogenesis of atherosclerosis in patients undergoing HD include diabetes, hypertension,
dyslipidemia, smoking, oxidative stress, inflammation and endothelial dysfunction. Since atherosclerosis has a long
subclinical phase it is important that pathogenic processes are identified while disease is still asymptomatic. Increased
carotid intima-media thickness (cIMT) is well recognized as an early indicator of subclinical atherosclerosis both in
the general population and in HD patients and it is associated with traditional and emerging CV risk factors. Several
studies have shown that carotid atherosclerosis is associated with circulating concentrations of soluble adhesion
molecules (CAMs) and omentin-1 protein. The aims of this study were to: a) investigate association of sICAM-1,
sVCAM-1, omentin-1 and other non-traditional risk factors with subclinical atherosclerosis; b) examine the diagnostic
value of these specific markers in the early detection of subclinical atherosclerosis and c) examine their role as
predictors of mortality in group of patients with subclinical atherosclerosis on regular HD.
Starting from November 2011, a cohort of 210 HD patients participated in this three-year follow-up study. The
subjects were divided into three groups according to the presence of atherosclerosis. Atherosclerotic disease was
assessed by measuring carotid intima-media thickness (IMT). Samplings were withdrawn at baseline and thereafter
every 12 months until the end of follow-up. Echocardiography was performed at baseline and at the end of follow-up.
IMT showed weak correlation with sICAM-1 (r=0.39, P=0.001), sVCAM-1 (r=0.27, P=0.015), and omentin-1 (r=-
0.25, P=0.020), also omentin-1 showed good correlation with parameters of systolic and diastolic function (r=0.52,
P=0.001 and r=0.51, P=0.001). Multivariate analysis showed that sICAM-1 and sVCAM-1 concentrations were a
strong independent correlate of IMT (P=0.031 and P=0.010, respectively). The Cox proportional analysis showed that
sICAM-1 and omentin-1 concentrations were strong predictors of cardiovascular death (HR=1.85, CI=1.18-2.32,
P=0.021 and HR=4.14, CI=1.38-12.1, P=0.004, respectively) and that serial measurements of these markers predict
IMT progression (HR=1.98, 95% CI=1.21-2.38, P<0.002 and HR=2.91, 95% CI=1.57-4.72, P<0.001, respectively).
Our study demonstrated that sICAM-1 and omentin-1 levels are strong predictors of cardiovascular death in HD
patients with subclinical atherosclerosis. . We believe that an increase in sICAM-1 and decrease in omentin-1 levels
could play an important role in the early progression of atherosclerosis.
Dentalna erozija danas je najčešća nekarijesna promjena na zubima, a nastaje zbog dugotrajnog djelovanja kiselih kemijskih čimbenika na površinu zuba, bez utjecaja bakterija. Ti se čimbenici dijele ...na unutarnje i vanjske. Unutarnji čimbenik je kiseli želučani sadržaj koji se zbog povraćanja može naći u usnoj šupljini, a vanjske čimbenike predstavljaju sve kisele tvari koje se unose u usta, najčešće pojedine vrste hrane i pića te lijekovi. Svrha ovog prikaza je upozoriti na mogući utjecaj kombiniranog lijeka s analgetičkim učinkom (aktivne komponente – paracetamol, propifenazon, kodein-fosfat i kofein) na promjene na zubima. U radu se opisuju vrlo specifične erozije na usnenim dijelovima zuba te njihova ljubičasto-plavkasta boja uočena tijekom obdukcije pothranjenog 35-godišnjeg muškarca koji je umro tijekom prijma na bolničko liječenje. Heteroanamnestički je dobiven podatak o dugogodišnjoj ovisnosti o kombiniranom analgetiku. Osim promjena na zubima, obdukcijom su ustanovljene erozije sluznice želuca, kronični vrijed u području dvanaesnika, kronična upala jetara te hipoplazija koštane srži. Uzrok smrti bila je obostrana gnojna upala pluća s velikim gangrenoznim žarištem lijevog pluća. Kemijskotoksikološkom analizom potvrđena je prisutnost svih farmakološki aktivnih tvari u mokraći, žučnoj tekućini, tkivu jetara i bubrega, želučanom sadržaju, te u ekstraktu zuba. Raspravlja se o vjerojatnom mehanizmu nastanka promjena na zubima i promjenama na drugim tkivima i organima, kao posljedicama dugotrajne ovisnosti o kombiniranom analgetiku. Ističe se važnost zuba kao prikladnoga za toksikološku analizu metodom plinske kromatografije te važnost interdiciplinarnog pristupa u rješavanju forenzičnih slučajeva. Posebno se ističe izolacija analgetika iz zuba, kao korisna dijagnostička metoda u slučaju sumnje na zloporabu tih lijekova.
Glycans constitute the most abundant and diverse form of the post-translational modifications. While genes
unequivocally determine the structure of each polypeptide, there is no genetic template for ...the glycan part.
Instead, hundreds of genes and their products interact in the very complex pathway of glycan biosynthesis
which is further complicated by environmental influences. Therefore, the aim of this thesis was to
determine the extent to which individual differences in immunoglobulin G and total plasma proteins
glycosylation patterns reflect genetic versus environmental influences. A twin study design was used and
study subjects were twins enrolled in the TwinsUK registry, a national register of adult twins. More than
4500 samples were analyzed by HILIC-UPLC (Hydrophilic Interaction Ultra Performance Liquid
Chromatography). A high contribution of the genetic component to N-glycome composition was found.
Variation in levels of 51 of the 76 IgG glycan traits studied was at least 50% heritable and only a small
proportion of N-glycan traits had a low genetic contribution. Heritability of plasma N-glycome was also
high, with half of the plasma glycan traits being at least 50% heritable. Further, epigenome-wide association
(EWA) analysis showed that methylation levels at some genes are also implicated in glycome composition,
both in those with high heritability and those with a lower genetic contribution. The study to investigate the
potential role of glycosylation in kidney function was also conducted. Fourteen IgG glycan traits were
associated with renal function in discovery population and remained significant after validation in an
independent subset of monozygotic twins discordant for renal disease, reflecting difference in
galactosylation, sialylation, and level of bisecting N-acetylglucosamine. Using the weighted correlation
network analysis (WGCNA) for IgG glycan traits, a correlation between low back pain (LBP) and glycan
modules was established. There was a weak positive correlation between pain phenotypes and "proantibody-
dependent cell-mediated cytotoxicity (ADCC)" WGCNA glycan modules (high bisecting Nacetylglucosamine
and low core fucose) and a weak negative correlation between pain phenotypes and
"anti-ADCC" module (high core fucose, no bisecting N-acetylglucosamine). This suggests that glycans are
promising candidates for biomarkers in many different diseases.
Glycans constitute the most abundant and diverse form of the post-translational modifications. While genes
unequivocally determine the structure of each polypeptide, there is no genetic template for the glycan part.
Instead, hundreds of genes and their products interact in the very complex pathway of glycan biosynthesis
which is further complicated by environmental influences. Therefore, the aim of this thesis was to
determine the extent to which individual differences in immunoglobulin G and total plasma proteins
glycosylation patterns reflect genetic versus environmental influences. A twin study design was used and
study subjects were twins enrolled in the TwinsUK registry, a national register of adult twins. More than
4500 samples were analyzed by HILIC-UPLC (Hydrophilic Interaction Ultra Performance Liquid
Chromatography). A high contribution of the genetic component to N-glycome composition was found.
Variation in levels of 51 of the 76 IgG glycan traits studied was at least 50% heritable and only a small
proportion of N-glycan traits had a low genetic contribution. Heritability of plasma N-glycome was also
high, with half of the plasma glycan traits being at least 50% heritable. Further, epigenome-wide association
(EWA) analysis showed that methylation levels at some genes are also implicated in glycome composition,
both in those with high heritability and those with a lower genetic contribution. The study to investigate the
potential role of glycosylation in kidney function was also conducted. Fourteen IgG glycan traits were
associated with renal function in discovery population and remained significant after validation in an
independent subset of monozygotic twins discordant for renal disease, reflecting difference in
galactosylation, sialylation, and level of bisecting N-acetylglucosamine. Using the weighted correlation
network analysis (WGCNA) for IgG glycan traits, a correlation between low back pain (LBP) and glycan
modules was established. There was a weak positive correlation between pain phenotypes and "proantibody-
dependent cell-mediated cytotoxicity (ADCC)" WGCNA glycan modules (high bisecting Nacetylglucosamine
and low core fucose) and a weak negative correlation between pain phenotypes and
"anti-ADCC" module (high core fucose, no bisecting N-acetylglucosamine). This suggests that glycans are
promising candidates for biomarkers in many different diseases.
Glikani predstavljaju najzastupljeniji i najraznolikiji oblik posttranslacijske modifikacije. Dok geni
nedvosmisleno određuju strukturu svakog polipeptida, za sintezu glikana ne postoji genski predložak.
Umjesto toga, stotine gena i njihovih produkata sudjeluju u vrlo kompleksnoj biosintezi glikana koju
okolišni utjecaji čine još složenijom. Stoga je cilj ovog doktorskog rada odrediti razmjer kojim genski i
okolišni čimbenici utječu na N-glikane imunoglobulina G (IgG) i ukupnih glikoproteina plazme. Da bi se
postigao navedeni cilj, upotrijebljena je studija na blizancima. Ispitanici su regrutirani TwinsUK registrom,
najvećim registrom blizanaca u Velikoj Britaniji. Više od 4500 uzoraka analizirano je HILIC-UPLC
metodom (kromatografijom vrlo visoke djelotvornosti temeljenoj na hidrofilnim interakcijama, eng.
Hydrophilic Interaction Ultra Performance Liquid Chromatography). Velika genska komponenta
(heritabilnost ≥ 50%) pokazana je za 51 od 76 glikanskih svojstava IgG-a. Nasuprot tome, samo je 12
glikanskih svojstava IgG-a pokazalo malu gensku komponentu. Heritabilnost plazmatskog N-glikoma
također se pokazala velikom. Polovica plazmatskih glikanskih svojstava bila je barem 50% heritabilna.
Epigenomska asocijacijska analiza pokazala je da razina metilacije na nekim genima također utječe na
sastav glikoma, i to na visokoheritabilne i niskoheritabilne glikane. Također je provedeno prvo istraživanje
potencijalne uloge IgG glikozilacije u funkciji bubrega. U prvoj je analizi pronađena značajna povezanost s
bubrežnom funkcijom za 14 glikanskih svojstava, a ostala je značajna i nakon validacije na neovisnoj
podskupini monozigotnih blizanaca diskordantnih za bubrežnu funkciju. Ta glikanska svojstva pripadaju
trima glavnim glikozilacijskim karakteristikama IgG-a: galaktozilaciji, sijalinizaciji i razini račvajućeg Nacetilglukozamina.
Koristeći se WGCNA (eng. weighted correlation network analysis) metodologijom,
provedena je mrežna analiza razina IgG glikana kod blizanaca, da bi se uspostavili klasteri koreliranih
glikana. Pronađene su povezanosti između tih klastera i fenotipova boli kod blizanaca s LBP-om. Opažena
je pozitivna korelacija između fenotipova boli i „pro-stanična citotoksičnost ovisna o protutijelima (eng.
antibody-dependent cell-mediated cytotoxicity, ADCC)“ WGCNA glikanskog modula (visoka razina
račvajućeg N-acetilglukozamina i niska razina sržne fukoze) te negativna korelacija između fenotipova boli
i „anti-ADCC“ modula (visoka razina sržne fukoze, bez račvajućeg N-acetilglukozamina). Rezultati
pokazuju da su glikani obećavajući biomarkeri za mnoge bolesti.
Moždani udar (MU) u djece heterogeni je poremećaj s višestrukom etiologijom, koja je još uvijek nepoznata u
približno 30% slučajeva. Genetički čimbenici nisu do kraja karakterizirani, a raspodjela ...gena kandidata za
nastanak MU različita je u pojedinim populacijama. U istraživanje je bilo uključeno 100 djece starosne dobi do
18 godina s potvrđenim MU i 100 zdrave djece podudarne po dobi i spolu djeci s MU. Napravljena je
genotipizacija 14 polimorfizama u 12 gena kandidata koji kodiraju proteine uključene u sustav zgrušavanja i
fibrinolize (FV Leiden, FV HR2, FII G20210A, beta-fibrinogen -455G>A, FXIII-A Val34Leu, PAI-1 4G/5G),
ljudskih trombocitnih antigena (HPA-1, -2, -3 i -5), metabolizma homocisteina (MTHFR C677T, MTHFR
A1298C) i intermedijernih rizicnih cimbenika (ACE I/D i ApoE 2-4). Utvrđena je cešća pojavnost MU u
dječaka, koja je uočena i u svim ispitivanim podskupinama. Najveći rizik za nastanak MU zabilježen je u prvoj
godini života, kao i 2,7 puta veći broj arterijskih ishemijskih MU (AIMU) u odnosu na hemoragijski MU
(HMU). Rezultati genotipizacije potvrdili su prije opisanu povezanost FV Leiden s nastankom (pojavom) MU.
Dokazana je i povezanost 4 polimorfizma (FV Leiden, FXIII-A Val34Leu, HPA-3 i apoE _2-4) s AIMU i to
različitih polimorfizama u pojedinim podskupinama prema spolu i dobi pojave AIMU, kao i povezanost
kombinacija genotipova polimorfizama MTHFR C677T i MTHFR A1298 (CC/AC) i dvostrukog heterozigotnog
oblika (GA/AG) polimorfizama FV Leiden i FV HR2 s AIMU, te kombinacije genotipova GA/AA s
perinatalnim AIMU. Utvrđen je i umjereno povećan rizik za pojavu HMU u nosioca haplotipa HA2 (HPA-
1a2a3a-ACED). Ovo istraživanje ukazalo je na povezanost pojedinih dosad nedovoljno istraženih polimorfizama
kao i kombinacija polimorfizama s etiologijom MU u djece u Hrvatskoj, što upotpunjuje i proširuje dosadašnje
spoznaje o etiološkim rizičnim čimbenicima.
Although stroke in children is a relatively rare and heterogeneous disorder with a wide range of identified risk
factors, the etiology of stroke is still undetermined in up to 30% of children. Taking into consideration that
genetic risk factors are incompletely characterized at present and the frequency of genetic factors may vary
among different populations, a genotype analysis was perforrmed of 14 polymorphisms in 12 candidate genes
encoding proteins of the coagulation and fibrinolysis systems (FV Leiden, FV HR2, FII G20210A, beta-fibrinogen -
455G>A, FXIII-A Val34Leu, PAI-1 4G/5G), human platelet alloantigens (HPA-1, -2, -3 and -5), homocysteine
metabolism (MTHFR C677T, MTHFR A1298C) and intermediate risk factors (ACE I/D and apoE 2-4) was
performed. The subject group comprised 100 children with a confirmed diagnosis of stroke aged £18 years and
100 age- and sex-matched control subjects. Obtained results were also analyzed in gender-specific stroke group
and subgroups according to the type of stroke: arterial ischemic (AIS) and hemorrhagic stroke (HS), and
according to the time of stroke onset (perinatal and childhood AIS). The predominance of boys was found
among (in) children with stroke and in all tested subgroups. The greatest risk for stroke was identified in the first
year of life, and AIS was found 2.7 times more frequently than HS. This case-control study has confirmed the
association between FV Leiden and stroke that was also observed in numerous studies so far, but it has also
shown that other previously not reported polymorhisms (FXIII-A Val34Leu, HPA-3 and the combination of FV
Leiden and FV HR2 polymorphisms) can be related to AIS in Croatian population. Analysis performed in
gender-specific stroke subgroups revealed the association of different polymorphisms in boys (FXII-A
Val34Leu, HPA-3 and combination of MTHFR polymorphisms) and girls (apoE 4 allele). The strongest
association (with OR>10.0) was found between FV Leiden and perinatal AIS for both genders whereas the
lowest risk for perinatal AIS was observed in girls who were carriers of at least one HPA-3b allele. Obtained
results have at least partially elucidated the role (impact) of new evaluated polymorphisms in the etiology of
stroke in children and their impact according to gender, type of stroke and time of stroke onset.
U okviru ovog doktorskog rada po prvi put je provedeno opsežno usporedno istraživanje fitokemijskog sastava i bioloških učinaka odabranih vrsta roda Thymus hrvatske flore (T. longicaulis C. Presl, T. ...praecox Opiz subsp. polytrichus (A. Kerner ex Borbás) Jalas, T. pulegioides L., T. serpyllum L. subsp. serpyllum i T. striatus Vahl). Kvalitativna i kvantitativna analiza biološki aktivnih sastavnica provedena je primjenom kromatografskih tehnika (TLC, RP-HPLC-DAD, LC-ESI-MS/MS i GC-MS) i spektrofotometrijskih metoda. Ružmarinska kiselina je bila najzastupljenija polifenolna sastavnica svih istraživanih vrsta. Većina
identificiranih flavonoida bila je iz skupine flavona. Najveći broj pripadao je derivatima luteolina, dok se udjelom isticao skutelarin. Istraživane Thymus vrste značajno su se razlikovale prema sadržaju i sastavu eteričnog ulja. Vrste T. longicaulis i T. pulegioides bile su bogate timolom i/ili karvakrolom, dok je 1,8-cineol bio glavna sastavnica vrste T. serpyllum subsp. serpyllum. U eteričnom ulju planinskih vrsta T. praecox subsp. polytrichus i T. striatus prevladavali su seskviterpenski spojevi. Biološki učinci eteričnih ulja, biljnih ekstrakata i fenolnih sastavnica ispitani su u prikladnim in vitro eksperimentalnim sustavima.
Eterična ulja odabranih Thymus vrsta inhibirala su rast bakterija S. aureus, E. faecalis i E. coli. Ustanovljeno je da se uklapanjem eteričnih ulja u liposome zadržava isti ili se može postići bolji antibakterijski učinak. Istraživanim biljnim ekstraktima dokazana su snažna antioksidacijska svojstva temeljena na sposobnosti hvatanja slobodnih radikala, redukciji i keliranju iona željeza te sposobnost inhibicije lipidne peroksidacije. Thymus ekstrakti su pokazali sposobnost inhibicije Src tirozin kinaze te su
smanjili produkciju proupalnog citokina IL-6 u splenocitima Balb/C miševa. Rezultati provedenih fitokemijskih i bioloških istraživanja pokazali su da, među Thymus vrstama koje rastu u Hrvatskoj, tankolisna majčina dušica (T. longicaulis) ima najveći potencijal za primjenu u farmaciji, usporediv s farmakopejskom vrstom T. vulgaris.
In this doctoral thesis, a comprehensive comparative study of the phytochemical composition and biological effects of selected Thymus species (T. longicaulis C. Presl, T. praecox Opiz subsp. polytrichus (A. Kerner ex Borbás) Jalas, T. pulegioides L., T. serpyllum L. subsp. serpyllum i T. striatus Vahl) from Croatian flora was conducted for the first time. Qualitative and quantitative chemical analyses of bioactive compounds were carried out using various chromatographic (TLC, RP-HPLC-DAD, LC-ESI-MS/MS and GC-MS) and spectrophotometric methods. Rosmarinic acid was found to be the most abundant polyphenolic constituent of all investigated species. Most of the identified flavonoids were from the group of flavones, the largest number belonged to luteolin derivatives, while scutellarin was present with the highest concentration. The investigated Thymus species were significantly different according to the content and composition of
essential oils. Thymol and/or carvacrol were found to be the main components of essential oils of T. longicaulis and T. pulegioides, while T. serpyllum subsp. serpyllum was rich in 1,8-cineole. The mountain species T. praecox subsp. polytrichus and T. striatus were characterised by a high percentage of sesquiterpenes. The biological effects of essential oils, plant extracts and phenolic constituents were studied using different in vitro assays. Selected Thymus essential oils inhibited growth of bacterial strains S. aureus, E. faecalis and E. coli. It was found that encapsulation of essential oils in liposomes can maintain or even improve their antibacterial efficiency. The studied plant extracts displayed strong antioxidant properties by acting as free radical scavengers and reducing agents and, additionally, by chelating transition metals and inhibiting lipid peroxidation. The Thymus extracts showed Src tyrosine kinase inhibitory activity and they
reduced the production of proinflammatory cytokine IL-6 in Balb/C mice splenocytes. The results of phytochemical and biological investigations suggest that, among Croatian Thymus species, T. longicaulis has the greatest potential for pharmaceutical usage comparable to that of pharmacopoeial species T. vulgaris.
Osteoartritis (OA) je najčešća bolest zglobova u razvijenim zemljama i vodeći je uzrok
kronične onesposobljenosti, napose ukoliko je bolešću zahvaćeno koljeno ili kuk. Kako uzrok
OA nije poznat, ...liječenje je prvenstveno usmjereno na uklanjanje simptoma i sprječavanje
funkcijske nesposobnosti. Od nefarmakoloških terapijskih postupaka čvrsti dokazi o
djelotvornosti postoje samo za kineziterapiju, no još uvijek nije utvrđena optimalna "doza"
(frekvencija, intenzitet i trajanje) vježbanja kao niti najučinkovitija vrsta vježbi. U kliničkoj
praksi se često primjenjuje i elektromagnetoterapija (EMT), o čijoj učinkovitosti u literaturi
postoje kontradiktorni podatci.
Cilj ovog istraživanja bio je usporediti učinkovitost osmišljenog kineziterapijskog programa i
njegove kombinacije s EMT na bol i funkcijsku sposobnost bolesnika s OA koljena.
U istraživanju su sudjelovala 92 ispitanika, metodom slučajnog izbora podijeljena u dvije
skupine: jednu, koja je provodila kineziterapijski program i drugu, u kojoj je uz
kineziterapijski program bila primijenjena i EMT.
Rezultati su pokazali da je u obim skupinama nastupilo statistički značajno smanjenje
intenziteta boli i osjećaja zakočenosti, poboljšanje bolesti mjereno globalnom ocjenom
liječnika i poboljšanje funkcijske sposobnosti, mjereno "Timed up and go" (TUG) testom i
WOMAC (od engl. Western Ontario and McMaster Universities Index of Osteoarthritis)
upitnikom. Ispitanici koji su samo provodili vježbe naveli su i značajno poboljšanje bolesti
prema vlastitoj globalnoj ocjeni, dok se u ispitanika koji su uz vježbe primali i EMT značajno
povećao opseg fleksije bolnog koljena.
Usporedbom skupina ispitanika pokazano je da nema statistički značajno većeg terapijskog
učinka u ispitanika koji su dodatno primali EMT u odnosu na one koji su samo provodili
kineziterapijski program, a poboljšanja u većini mjerenih varijabli bila su i izraženija u
skupini ispitanika bez dodatne primjene EMT.
Zaključno, u ovom je istraživanju dokazan pozitivan terapijski učinak osmišljenog
kineziterapijskog programa na bol i funkcijsku sposobnost bolesnika s OA koljena, ali ne i
hipoteza prema kojoj će kombinacija kineziterapije i EMT imati bolji učinak na iste parametre
u odnosu na primjenu samo kineziterapije.
Rezultati istraživanja mogu značajno utjecati na svakodnevni klinički rad i općenito na
liječenje i rehabilitaciju bolesnika s OA koljena.
Osteoarthritis (OA) is the most common joint disease and a leading cause of chronic
disability, in large part due to knee or hip involvement. Etiology is unknown, so the treatment
is focused on symptoms elimination and functional disability prevention. Among
nonpharmacologic modalities, only the kinesiotherapy effectiveness was confirmed while
neither the optimal exercise „dosage“ (frequency, intensity and duration) nor the most
effective type of exercise have been defined. Pulsed electromagnetic fields (PEMF) are
frequently used in the treatment, although the literature provides contradictory data related to
its effectiveness.
The aim of this research was to compare the effect of designed kinesiotherapy program and its
combination with PEMF on pain and functional ability in patients with knee OA.
Ninety-two patients were randomly divided into two groups: in the first one the patients
underwent kinesiotherapy program and in the second, besides that program, patients were
treated with PEMF.
In both groups results showed statistically significant reduction of pain intensity and stiffness,
disease improvement measured by physician’s global assessment and improvement of
functional ability measured by "Timed up and go" (TUG) test and WOMAC (Western Ontario
and McMaster Universities Index of Osteoarthritis) questionnaire. Additionally, the patients
that performed exercises only, declared statistically significant disease improvement
according to their global assessment, while the patients that received PEMF therapy after the
identical exercise program had statistically significant improvement of painful knee flexion.
Comparison of the patient’s groups didn’t show any statistically significant improvements in
therapeutic effect in group that combined exercises and PEMF therapy. On the contrary,
increase in most measured variables was even emphasized in group without additional PEMF
therapy.
In conclusion, this research proved the positive therapeutic effect of clearly designed
kinesiotherapy program on pain and function ability in patients with knee OA, while the
hypothesis about better parameters indicators in group who underwent combined therapy
(kinesiotherapy and PEMF) was not substantiated.
The results of the research may be important for daily clinical practice in treatment and
rehabilitation of patients with knee OA.
Osteoartritis (OA) je najčešća bolest zglobova u razvijenim zemljama i vodeći je uzrok
kronične onesposobljenosti, napose ukoliko je bolešću zahvaćeno koljeno ili kuk. Kako uzrok
OA nije poznat, liječenje je prvenstveno usmjereno na uklanjanje simptoma i sprječavanje
funkcijske nesposobnosti. Od nefarmakoloških terapijskih postupaka čvrsti dokazi o
djelotvornosti postoje samo za kineziterapiju, no još uvijek nije utvrđena optimalna "doza"
(frekvencija, intenzitet i trajanje) vježbanja kao niti najučinkovitija vrsta vježbi. U kliničkoj
praksi se često primjenjuje i elektromagnetoterapija (EMT), o čijoj učinkovitosti u literaturi
postoje kontradiktorni podatci.
Cilj ovog istraživanja bio je usporediti učinkovitost osmišljenog kineziterapijskog programa i
njegove kombinacije s EMT na bol i funkcijsku sposobnost bolesnika s OA koljena.
U istraživanju su sudjelovala 92 ispitanika, metodom slučajnog izbora podijeljena u dvije
skupine: jednu, koja je provodila kineziterapijski program i drugu, u kojoj je uz
kineziterapijski program bila primijenjena i EMT.
Rezultati su pokazali da je u obim skupinama nastupilo statistički značajno smanjenje
intenziteta boli i osjećaja zakočenosti, poboljšanje bolesti mjereno globalnom ocjenom
liječnika i poboljšanje funkcijske sposobnosti, mjereno "Timed up and go" (TUG) testom i
WOMAC (od engl. Western Ontario and McMaster Universities Index of Osteoarthritis)
upitnikom. Ispitanici koji su samo provodili vježbe naveli su i značajno poboljšanje bolesti
prema vlastitoj globalnoj ocjeni, dok se u ispitanika koji su uz vježbe primali i EMT značajno
povećao opseg fleksije bolnog koljena.
Usporedbom skupina ispitanika pokazano je da nema statistički značajno većeg terapijskog
učinka u ispitanika koji su dodatno primali EMT u odnosu na one koji su samo provodili
kineziterapijski program, a poboljšanja u većini mjerenih varijabli bila su i izraženija u
skupini ispitanika bez dodatne primjene EMT.
Zaključno, u ovom je istraživanju dokazan pozitivan terapijski učinak osmišljenog
kineziterapijskog programa na bol i funkcijsku sposobnost bolesnika s OA koljena, ali ne i
hipoteza prema kojoj će kombinacija kineziterapije i EMT imati bolji učinak na iste parametre
u odnosu na primjenu samo kineziterapije.
Rezultati istraživanja mogu značajno utjecati na svakodnevni klinički rad i općenito na
liječenje i rehabilitaciju bolesnika s OA koljena.
Svrha je istraživanja bila odrediti fizičke i kemijske karakteristike terapijskog polja
visokofrekvencijskog generatora ozona.
U terapijskom polju je izmjeren spektar plazma elektrode u području ...540-940 nm (atomske
linije Ne I i O I) dok se kod izboja kroz zrak bilježi i spektar mikro-izboja u području
290-440 nm (molekularne linije N2 i N2+). Na uzorku tkiva su izmjerene vrijednosti
efektivnog napona (22-95 mV), i efektivne struje (0,22-0,95 mA) na osnovu kojih su
procijenjene vrijednosti gustoće struje (4-19 μA/mm2) kao i snage (5-65 μW) te predane
energije u ovisnosti o vremenu izlaganja (0,14-10,75 mJ kod trajanja zahvata 30-
120 sekundi). Kod oba modaliteta primjene, u frekvencijskom području 33 kHz, zabilježena je
jakost magnetskog polja (1-3,07 A/m) i gustoće magnetskog toka (1,25-3,86 μT). Prilikom
pojave mikro-izboja u zračnom međuprostoru, u frekvencijskom području iznad 40 MHz, se
bilježi magnetsko polje jakosti (5-49 mA/m) te gustoće magnetskog toka od (6,5- 62,7 nT).
Jakost električnog polja na udaljenosti od 10 mm od elektrode iznosi 248-586 V/m. Tijekom
provedbe zahvata zabilježene su značajne promjene u temperaturi površine tkiva te one
ovisno o intenzitetu i trajanju zahvata (30-120 sek) iznose od 2.6 do 12,2°C. Statistička
analiza rezultata ukazala je na značajnu razliku u vrijednostima fizičkih veličinama s obzirom
na dva modaliteta primjene.
Na osnovu navedenih dokaza se zaključuje da djelovanje ispitivanog uređaja ne bi trebalo
pripisivati isključivo ozonu, kao što je to dosad bio slučaj, već kumulativnom učinku svih,
ovim istraživanjem dokazanih, fizičkih i kemijskih čimbenika koji su tijekom zahvata prisutni
u terapijskom polju.
A high-frequency ozone generator Ozonix (Biozonix GmbH, Germany) is classified as a
medical ozone generator, thus its therapeutic effects have, so far, been attributed exclusively
to ozone effect. The operative principle of the device and ozone production through a
dielectric barrier discharge using glass electrodes filled with a noble gas requires presence of
additional physical and chemical factors, besides ozone, in the treatment field during the
procedure. The purpose of the study was to determine the physical and chemical properties in
the treatment field of the said device. For that purpose, measuring of emission spectra, electric
and magnetic values as well as changes in surface temperature on tissue samples, as functions
of the selected intensity and exposure time, was conducted, and two basic application
modalities (contact discharge and air gap discharge) were used.
Materials and Methods: Optical emission spectroscopy was used to record the spectrum of a
plasma electrode’s glow discharge, a micro discharge occurring in the aerial interspace (air
gap) between the electrode and the targeted surface and the spectrum of the discharge through
a liquid medium on the surface of the tissue sample. By measuring electromagnetic values
present in the treatment field, physical values of electric and magnetic fields were recorded, as
well as currents passing through the treatment field. Thermographic measurements were used
to record temperature changes at different intensities (10-100%) and exposure durations (30,
60, 90 and 120 seconds).
Results: In the treatment field, two main emission spectra were recorded. Spectral lines of
plasma electrode’s glow discharge were recorded in the optical range between 540 and
940 nm, which were identified as atomic lines of neon (Ne I) and oxygen atoms (O I). This
spectrum is observed in all treatment modalities. During the formation of micro-discharges in
the air gap between the electrode and the treated surface, spectral lines were recorded in the
optical range between 290 and 440 nm, which were identified as molecular lines of N2 and N2+.
On the tissue samples (pork skin), as a function of output intensity (10-100%), electrical
values were recorded by use of oscilloscope and low impedance (Uin = 100) probe: RMS
voltage (22-95 mV), RMS current (0,22- 0.95 mA). These values were used to calculate
current density (4-19 μA/mm2) as well as power (5-65 μW). Using the same duration periods
as the ones used in the examination of temperature change at the tissue surface (30-120 sec),
the values of committed (delivered) energy were estimated (0,14-10,75 mJ).
Magnetic fields were recorded at two frequency ranges. In the frequency range of 33 kHz,
magnetic field was recorded in both treatment modalities as a function of intensity (10-100%)
for direct contact discharge H(1,93 ± 0,59 A/m), B (2,43 ± 0,75 μT), f (32151 ± 985Hz) and
air gap discharge H (2,5 ± 0,52 A/m), B (3,14 ± 0,65 μT), f (34526 ± 833 Hz). Only in the
case of micro-discharge formation in the air gap were additional peaks of magnetic field
observed in a wide frequency range from 40 to above 400 MHz. Due to the low intensity,
magnetic field values were calculated only for 41.5 MHz peak, where H field was
0,032 ± 0,014 A/m and B field was 40 ± 17,4 nT. At 10 mm distance from plasma electrode
surface, electric field strength was recorded in the range from 248 to 586 V/m depending on
the selected intensity (10-100%). During the treatment, a mild increase of temperature on the
surface of the tissue samples was recorded (2,6-12,2°C) as a function of intensity (20-100%)
and exposure time (30-120 seconds).
Conclusion: The results of the conducted research confirm the presence of mildly ionized
cold atmospheric plasma in the treatment field during air gap discharge. In both treatment
modalities, the presence of light in visible red and near infrared spectra, electric field,
magnetic fields, the currents flowing through the tissue and mild heat generated on the surface
of the tissue sample were confirmed in the treatment field. Furthermore, a significant
difference was determined in the values of physical properties with respect to the application
modality, where the values of temperature increase and magnetic field strengths are
significantly higher in the air gap modality whilst the values of electric current (and other
derived values) are lower compared to direct contact modality.
According to the results of this research, it can be concluded that treatment effects with the
tested devices must be attributed not only to ozone, but to cumulative effects of all physical
and chemical factors, the presence of which was recorded in the treatment field during this
research.
Svrha je istraživanja bila odrediti fizičke i kemijske karakteristike terapijskog polja
visokofrekvencijskog generatora ozona.
U terapijskom polju je izmjeren spektar plazma elektrode u području 540-940 nm (atomske
linije Ne I i O I) dok se kod izboja kroz zrak bilježi i spektar mikro-izboja u području
290-440 nm (molekularne linije N2 i N2+). Na uzorku tkiva su izmjerene vrijednosti
efektivnog napona (22-95 mV), i efektivne struje (0,22-0,95 mA) na osnovu kojih su
procijenjene vrijednosti gustoće struje (4-19 μA/mm2) kao i snage (5-65 μW) te predane
energije u ovisnosti o vremenu izlaganja (0,14-10,75 mJ kod trajanja zahvata 30-
120 sekundi). Kod oba modaliteta primjene, u frekvencijskom području 33 kHz, zabilježena je
jakost magnetskog polja (1-3,07 A/m) i gustoće magnetskog toka (1,25-3,86 μT). Prilikom
pojave mikro-izboja u zračnom međuprostoru, u frekvencijskom području iznad 40 MHz, se
bilježi magnetsko polje jakosti (5-49 mA/m) te gustoće magnetskog toka od (6,5- 62,7 nT).
Jakost električnog polja na udaljenosti od 10 mm od elektrode iznosi 248-586 V/m. Tijekom
provedbe zahvata zabilježene su značajne promjene u temperaturi površine tkiva te one
ovisno o intenzitetu i trajanju zahvata (30-120 sek) iznose od 2.6 do 12,2°C. Statistička
analiza rezultata ukazala je na značajnu razliku u vrijednostima fizičkih veličinama s obzirom
na dva modaliteta primjene.
Na osnovu navedenih dokaza se zaključuje da djelovanje ispitivanog uređaja ne bi trebalo
pripisivati isključivo ozonu, kao što je to dosad bio slučaj, već kumulativnom učinku svih,
ovim istraživanjem dokazanih, fizičkih i kemijskih čimbenika koji su tijekom zahvata prisutni
u terapijskom polju.
