PATHOMORPHOLOGICAL AND COMPUTERIZED TOMOGRAPHY STUDY OF IBUPROFEN AND COMBINATION WITH 2-PHENYL-3-CARBETHOXY-4-DIMETHYLAMINOMETHYL-5-OKSIBENZOFURAN HYDROCHLORIDE (VINBORON) INFLUENCE ON THE JOINTS OF LOWER EXTREMITIES OF RATS WITH EXPERIMENTAL RHEUMATOID ARTHRITIS

The most frequent adverse reactions to nonsteroidal anti-inflammatory drugs are upper and lower gastrointestinal tract disorders, hepatotoxicity, nephrotoxicity, a decrease in the synthesis of proteoglycans of the cartilaginous tissue and allergic reactions. In our previous studies we have found out that Ukrainian vinboron, the antispasmodic with polytrophic pharmacological effects, can improve the safety profile of the popular non-steroidal antiphlogistics ibuprofen and can also potentiate its anti-inflammatory effect. That fact has become the background to a detailed study of the therapeutic action mechanisms of these drugs when they are used together.

Aim of the study. To characterize the influence of vinboron (11 mg/kg, i.g.) on the anti-inflammatory activity of ibuprofen (218 mg/kg, i.g.) when combined, in a model of experimental rheumatoid arthritis according to pathomorphological, morphometric and cone-ray computerized tomography of rats’ hind limbs.

Materials and methods. To reproduce the conditions for the administration of ibuprofen corresponding to its clinical use, we selected a model of experimental rheumatoid arthritis-adjuvant arthritis (AA) in rats. On the 28^th  day the animals were excluded from the experiment. It was followed by the amputation of the hind limbs at the level of the hocks (art. coxae) and fixation of the material in 10.0% aqueous solution of neutral formalin. The ankle joints were decalcified. The drugs were prepared according to the standard procedure. Microscopy and photographing of histological preparations were carried out with the light microscope OLIMPUS BX 41 at magnifications of 40, 100, 200 and 400 times. The obtained images were processed and morphometry was carried out with the program “Quick PHOTO MICRO 2.3”. The contents of the cellular elements was determined on the basis of the equivalent unit area (1 mm2). Cone-ray computerized tomography was carried out with a digital panoramic cone-ray tomographic scanner “Point 3D Combi 500” (PointNix, South Korea), using a planar sensor with a scanning site of 9 × 12 mm and a radial load of 30-100 μSv. Scanning was carried out with a voltage of 52.0 kV and the electric current of 6.0 mA. For the integral characteristics of the linear dimensions of rats’ hind limbs joints we proposed a formula for calculating the joint volume (Patent of Ukraine No. UA 117538). For the integral characteristics of the dimensions of rats’ hind limbs linear joints we proposed a formula for calculating the joint volume (Patent of Ukraine No. UA 117538).

Results and discussion. The use of ibuprofen in combination with vinboron was accompanied by restoration of the structure and function of the cartilage and bone tissues, evidenced by the appearance of acidic and sulfated glycosaminoglycans. The restoration of the bone tissue was realized by the proliferation of the connective tissue and its transformation into cartilage. The synovial membrane was hyperplastic with proliferating synoviocytes of both macrophage and secretory types. In rare cases, the phenomenon of moderate edema and minor focal lymphohistiocytic infiltration persisted. Thus, the use of ibuprofen and ibuprofen in combination with vinboron in rats with adjuvant arthritis resulted in a reduction or absence of inflammation in both the joint and periarticular tissues. The analysis of the results of cone-ray computerized tomography and digital radiography showed that against the background of simultaneous administration of ibuprofen and vinboron there was a tendency to increase the anti-inflammatory effect of the studied antiphlogistics on the rats’ joints affected by adjuvant arthritis. On the 28-th day of the experiment there was the indicatation of decrease of the volume of the hind limb joints by an average of 33.8% compared to the control animals and a near-complete restoration of the width of the joint gaps of the joints examined. It was also determined that vinboron can harmonize the anti-inflammatory effect of ibuprofen when they are used together.

Сonclusion. The use of vinboron in combination with ibuprofen leads to a more pronounced pathogenetic anti-inflammatory structural-modifying effect of this antiphlogistics.

1. Mazurov VI. Klinicheskaya revmatologiya. Rukovodstvo dlya vrachej [Clinical rheumatology. Guide for doctors]. SPb.: publ. “Foliant”. 2005. 496 р. Russian.

2. Nasonov EL, Nason VA. Revmatologiya Nacionalnoe rukovodstvo [Rheumatology. National leadership]. Moscow: GEOTAR-Media. 2008. 737 p. Russian.

3. Raskin TA, Letaeva MV, Koroleva MV. Profil pobochnyh effektov nesteroidnyh protivovospalitelnyh preparatov pri revmaticheskih zabolevaniyah [A profile of the side effects of non-steroidal anti-inflammatory drugs in rheumatic diseases]. Modern rheumatology journal. 2011;4:53–7. DOI: 10.14412/1996-7012-2011-699. Russian.

4. Strygol SYu. Farmakologicheskie svojstva i problemy bezopasnosti primeneniya NPVP selektivnyh i specificheskih ingibitorov ciklooksigenazy-2 [Pharmacological properties and safety problems of NSAID use – selective and specific inhibitors of cyclooxygenase-2]. Provizor. 2005;2:37–42. Russian.

5. Shih AR, Misdraji J. Drug-induced pathology of the upper gastrointestinal tract. Diagnostic Histopathology. 2017;23(2):84–95. DOI: 10.1016/j.mpdhp.2017.03.002

6. Blackler RW, Gemici B, Manko A, Wallace JL. NSAID-gastroenteropathy: new aspects of pathogenesis and prevention. Curr. Opin. Pharmacol. 2014;11:11–6. DOI: 10.1016/j.coph.2014.05.008

7. Masso Gonzalez EL, Pastrignani P, Tacconelli S, Garcia Rodriguez LA. Variability among nonsteroidal antiinflammatory drugs in risk of upper gastrointestinal bleeding. Arthritis Rheum. 2010;62(6):1592–601. DOI: 10.1002/art.27412

8. Lanas A, Garcia-Rodriguez LA, Arroyo MT, Gomollon F, Feu F, Gonzalez-Perez A, Zapata E, Bastida G, Rodrigo L, Santolaria S, Guell M, de Argila CM, Quintero E, Borda F, Pique JM. Risk of upper gastrointestinal ulcer bleeding associated with selective cyclo-oxygenase-2 inhibitors, traditional non-aspirin non-steroidal anti-inflammatory drugs, aspirin and combinations. Gut. 2006;55:1731–38.

9. Evseev MA. Povrezhdenie kishechnoj trubki nesteroidnymi protivovospalitelnymi preparatami klinicheskoe znachenie patogenez vozmozhnosti profilaktiki [Damage to the intestinal tube by non-steroidal anti-inflammatory drugs: clinical significance, pathogenesis, prevention possibilities]. Neurology, neuropsychiatry, psychosomatics. 2013;1:79–87. DOI: 10.14412/2074-2711-2013-2405. Russian.

10. Hladkykh FV, Stepanyuk NG. Suchasni shlyahi poslablennya ulcerogennosti nesteroidnih protizapalnih zasobiv: dosyagnennya nevirishen pitannya ta shlyahi optimizacii [New approaches to reduce ulcerogenity of nonsteroidal anti-infl ammatory drugs: achievements, unsolved issues and ways to optimize]. Zaporozhskij medicinskij zhurnal. 2014;2(83):82–6. DOI: https: 10.14739/2310-1210.2014.2.25437. Ukrainian.

11. Hladkykh FV. Harakteristika mekhanizmov antiulcerogennogo dejstviya agonistov vaniloidnyh receptorov TRPV1 na modeli gastropatii inducirovannoj acetilsalicilovoj kislotoj [Characteristics of the mechanisms of antiulcerogenic action of vanilloid receptor agonists (TRPV1) on the model of gastroepathy induced by acetylsalicylic acid]. Pharmacy & Pharmacology. 2017;5(3):283–301. DOI: 10.19163/2307-9266-2017-5-3-283-301. Russian.

12. Hladkykh FV, Stepanyuk NG. Harakteristika protizapalnoi ta znebolyuyuchoi aktivnosti ibuprofenu ta jogo kombinacii z vinboronom na modeli adyuvantnogo artritu u shchuriv [Characterization of anti-inflammatory and analgesic activity of ibuprofen and its combination with vinoboron on the model of adjuvant arthritis in rats]. Bulletin of scientific research. 2015;2(79):108–11. DOI: 10.11603/2415-8798.2015.2.5617. Ukrainian.

13. Hladkykh FV, Stepanyuk NG, Stepanyuk GI, Sokirko MV. Patent of Ukraine to the utility model No. UA 107533 U MPK A61K 31/135 (2006.01). Zastosuvannya vinboronu dlya potenciyuvannya antiflogistichnogo efektu ibuprofenu [Application of vinoboron for potentiating the antiflologic effect of ibuprofen]. Applicant and patent holder Vinnytsia National Medical University. No. u. 2015 12613; stated. 12.2.2015; has published Jun 10, 2011, Bul. No. 11. Ukrainian.

14. Hladkykh FV, Stepanyuk NG., Student VO., Pogrebennik YYa., Yurkevich BYe. Ocinka efektivnosti likuvannya eksperimentalnogo revmatoidnogo artritu ibuprofenom ta jogo kombinaciyu z vinboronom za danimi konusno-promenevoi kompyuternoi tomografii ta cifrovio rentgenografii zadnih kincivok shchuriv [Estimation of the effectiveness of treatment of experimental rheumatoid arthritis with ibuprofen and its combination with vinoboron according to the data of cone-ray computer tomography and digital Х-ray of the hind limbs of rats]. Pharmacology and drug toxicology. 2017;1(52):12–24. Ukrainian.

15. Chernoyvan NG, Voloshchuk NI, Bukhtiarova TA, Shalamay AS, Kurilyuk AС. Vpliv vinboronu na velichinu protizapalnoi ta znebolyuyuchoi dii diklofenaku pri adyuvantnomu artriti [Effect of vinoboron on the amount of anti-inflammatory and analgesic activity of diclofenac in adjuvant arthritis]. Pharmacology and drug toxicology. 2008; 1–3:57–9. Ukrainian.

16. Hladkykh FV, Chovgan IS, Student VO, Polozov SD, Yurkevich BYe. Ocinka efektivnosti protizapalnoi terapii eksperimentalnogo revmatoidnogo artritu ibuprofenom ta jogo kombinaciyu z vinboronom za pokaznikami remodelyuvannya kistkovoi tkanini [Evaluation of the effectiveness of anti-inflammatory therapy of experimental rheumatoid arthritis ibuprofen and its combination with vinoboron on bone remodeling indicators]. Materials of the XIV International Scientific Conference of Students and Young Scientists “The First Step in Science-2017” (April 26–28, 2017): theses of additional. Vinnitsa. 2017:509. Ukrainian.

17. Pearson CM, Wood FD. Studies of polyarthritis and other lesions induced in rats by injection of mycobacterial adjuvant. I. General clinical and pathological characteristics and some modifying factors. Arthr. and Rheum. 1959;2:440-59. DOI: 10.1002/1529-0131(195910)2:5<440::AID-ART1780020510>3.0.CO;2-N

18. Stefanov OV. Doklinichni doslidzhennya likarskih zasobiv: metod. rekomend. [Preclinical research of medicinal products: method. recommend] K.: VD “Avicena”. 2001. 527 p. Ukrainian.

19. Khubriev RU. Rukovodstvo po eksperimentalnomu doklinicheskomu izucheniyu novyh farmakologicheskih veshchestv [Manual on experimental (preclinical) study of new pharmacological substances]. Moscow. 2005:695–709. Russian.

20. Freund J. Some aspects of active immunization. Annual Review of Microbiology. 1947;1:291-308.

21. Newbould B.B. Chemotherapy of arthritis induced in rats by mycobacterial adjuvant. Br. J. Pharmacol. Chemo-ther. 1963;21(1):127–36.

22. Saratikov AS, Wengerovsky AI, Prischept TP. Adyuvantnaya bolezn (morfologiya, patogenez, eksperimentalnaya terapiya): monografiya [Adjuvant disease (morphology, pathogenesis, experimental therapy): monograph]. Tomsk: Izdat. Tomsk University. 1983. 101 p. Russian.

23. Snekhalatha U, Anburajan M, Venkatraman B, Menaka M. Evaluation of complete Freund’s adjuvant-induced arthritis in a Wistar rat model. Comparison of thermography and histopathology. Zeitschrift fur Rheumatologie. 2013; 72(4):375–82. DOI: 10.1007/s00393-012-1083-8

24. Rybolovlev YuR, Rybolovlev RS. Dozirovanie veshchestv dlya mlekopitayushchih po konstantam biologicheskoj aktivnosti [Dosage of substances for mammals according to the constants of biological activity]. Reports of the Academy of Sciences of the USSR. 1979; 247(6):1513–6. Russian.

25. Rainsford KD. Ibuprofen: pharmacology, therapeutics and side effects: monograph. Heidelberg: Springer Basel. 2012. 259 р.

26. Hladkykh FV, Stepanyuk NG. Vinboron pershij ukrainskij gastroprotektor agonist vaniloidnih receptoriv (TRPV1) [Vinboron: The first Ukrainian gastroprotector – hvanilloid receptor agonist (TRPV1)]. Pharmacology and drug toxicology. 2016; 4–5(50):20–9. Ukrainian.

27. Grinev AN, Panischev EK, Stolyarchuk AA, Ivanova NI, Piskun RP. Novyj lekarstvennyj preparat fenikaberan [New medicinal product of fenicaberane]. Pharmaceutical Chemistry Journal. 1979;11:118-21. Russian.

28. Chekman IS, Stolyarchuk OO, Gayoviy VP. Eksperimentalno-klinichna harakteristika preparatu Fenikaberan [Experimental-clinic characteristics of the drug “Fenikaberan”]. Farmacevticheskij zhurnal. 1992;5–6:76–8. Ukrainian.

29. Avtandilov GG. Osnovy patologoanatomicheskoj praktiki. Rukovodstvo (izdanie trete dopolnennoe) [Fundamentals of pathoanatomical practice. Manual (third edition augmented)]. M: Russian Medical Academy of Postgraduate Education. 2007. 480 p. Russian.

30. Golofeevsky VYu, Shcherbak SG. Sochetannaya okraska gistologicheskih srezov osnovnym korichnevym i prochnym zelenym [Combined color of histological sections of the main brown and durable green]. Arhiv anatomii gistologii i embriologii. 1987;4:101. Russian.

31. Sapozhnikov AG, Dorosevich AE. Gistologicheskaya i mikroskopichekaya tekhnika: Rukovodstvo [Histology and microscopy technique: Manual]. Smolensk: SAU. 2000. 476 р. Russian.

32. Shubich MG. Metod elektivnoj okraski kislyh sulfatirovannyh mukopolisaharidov osnovnym korichnevym [Method of elective coloring of acid (sulfated) mucopolysaccharides by the main brown]. Bulletin of experimental biology and medicine. 1961; 2:116–20. Russian.

33. Bagriy MM, Dibrova VA, Popadinets OG, Grischuk MІ. Metodiki morfologichnih doslidzhen: monografiya [Methods of morphological doslands: monograph]. Vinnytsya: Nova kniga. 2016. 328 p. Ukrainian.

34. Aurell Y, Malac M, Forslind K. Work in progress: cone beam computerized tomography (CBCT or extremity CT) – a new tool for evaluation of erosions in patients with rheumatoid arthritis. European Society of Musculoskeletal Radiology. 2014. DOI: 10.1594/essr2014/P-0028

35. Al-Ekrish AA, Ekram MA. A comparative study of the accuracy and reliability of multidetector computerized tomography and cone-beam computerized tomography in the assessment of dental implant site dimension. Dentomaxillofacial Radiollogy. 2011; 40:67–75. DOI: 10.1259/dmfr/27546065

36. Raskin TA, Ushakov AV. Ocenka sostoyaniya kostnoj tkani metodom kompyuternoj tomografii u bolnyh revmatoidnym artritom [Evaluation of bone tissue by the method of computerized tomography in patients with rheumatoid arthritis]. Rheumatology Science and Practice. 2002;1:20–2. Russian.

37. Reutsky IA, Marinin VF, Glotov AV. Diagnostika revmaticheskih zabolevanij Rukovodstvo dlya vrachej [Diagnosis of rheumatic diseases. Manual for doctors]. M .: OOO “Med. inform. agenstvo”. 2011:260-87. Russian.

38. Van der Heijde D.M. Radiographic imaging: “gold standard” for assessment of disease progression in rheumatoid arthritis. Rheumatology. 2000;39:9–16. DOI: 10.1038/35046102

39. Hladkykh FV, Stepanyuk NG, Student VO, Stepanyuk GI, Sokirko MV. Patent of Ukraine for Utility Model No. UA 117538 U MPK (2017.01), A61K 31/135 (2006.01 A61P 29/00 Sposib ocinki protizapalnoi aktivnosti farmakologichnih rechovin za velichinoyu ob mu dribnih suglobiv zadnih kincivok u shchuriv na modeli revmatoidnogo artritu [A method for evaluating the anti-inflammatory activity of pharmacologic substances by the size of the volume of small joints of the hind limbs in rats on the model of rheumatoid arthritis]. Applicant and patent holder Vinnitsa National Medical University. No. 2017 01095; stated. 02/06/2017; Published 26/06/2017, Byul. No. 12. Ukrainian.

40. Reznikov OG. Zagalni etichni principi eksperimentiv na tvarinah [General ethical principles of experiments on animals]. Endocrinology. 2003;8(1):142–5. Ukrainian.

41. Zar JH. Biostatistical analysis (5 ed.). Prentice-Hall, Englewood. 2014.