Резюме
Бутко Я.О. Дослідження лікувальної дії препаратів з глюкокортикостероїдами та керамідами в умовах альтеративного запалення.
Проведено вивчення антиальтеративної дії крему «Мометазон з керамідами» та мазі «Метилпреднізолон з керамідами» на моделі асептичного запалення шкіри у щурів. Результати дослідження показали, що розроблені препарати достовірно зменшують площу відкриття виразок у порівнянні з контрольною патологією та рівень антиальтеративної дії крему «Мометазон з керамідами» та мазі «Метилпреднізолон з керамідами» склав 74,4% та 76,7%, відповідно. При застосуванні препаратів з ГКС та керамідами відмічалась нормалізація системних показників крові у щурів, але інтактних значень вони не набули, що свідчить про зменшення потрапляння ГКС до кровообігу та зменшення ризику виникнення системних побічних реакцій ГКС. Отже, перспективним є подальше вивчення запропонованих комбінацій ГКС з керамідами з метою клінічного впровадження їх в дерматологію.
Ключові слова: глюкокортикостероїди, кераміди, запалення, побічні реакції.
Резюме
Бутко Я.А. Исследование лечебного действия препаратов с глюкокортикостероидами и керамидами в условиях альтеративного воспаления.
Проведено изучение антиальтеративного действия крема «Мометазон с керамидами» и мази «Метилпреднизолон с керамидами» на модели асептического воспаления кожи у крыс. Результаты исследования показали, что разработанные препараты достоверно уменьшают площадь вскрытия язв по сравнению с контрольной патологией и уровень антиальтеративного действия крема «Мометазон с керамидами» и мази «Метилпреднизолон с керамидами» составил 74,4% и 76,7%, соответственно. При применении препаратов с ГКС и керамидами отмечалась нормализация системных показателей крови у крыс, однако интактных значений они не приобрели, что свидетельствует о снижении всасывания ГКС в кровоток и уменьшении риска возникновения системных побочных реакций ГКС. Следовательно, перспективным является дальнейшее изучение предложенных комбинаций ГКС с керамидами с целью клинического внедрения их в дерматологию.
Ключевые слова: глюкокортикостероиды, керамиды, воспаление, побочные реакции.
Summary
Butko Y. The research of therapeutic effect of the drugs with corticosteroids and ceramides in the conditions of an alterative inflammation.
The study of antialterative action of cream “mometasone with ceramides” and ointment “Methylprednisolone with ceramides” on the model of aseptic inflammation of skin of rats was conducted. The results of research showed that the developed drugs significantly reduce the area of open ulcers in comparison with control pathology and level of antialterative effect of the cream “Mometasone with ceramides” and ointment “ Methylprednisolone with ceramides” made up 74.4% and 76.7%, respectively. At application of drugs with corticosteroids and ceramides normalization of systemic blood parameters of rats was observed, however they didn’t gain intact values, that testifies about decrease of absorption of GKS in a blood-groove and reduction of risk of emergence of system side effects of GKS. Therefore, further studying of the offered combinations of GKS with ceramides for the purpose of their clinical introduction in dermatology is perspective.
Key words: steroids, ceramides, inflammation, side effects.

Література
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UDC 615.015:615.454.122

THE research of therapeutic effect of THE drugs with corticosteroids and ceramides in the conditions of alterative  inflammation

Butko Y.A.

The National University of Pharmacy, Kharkiv, Ukraine

yaroslavabutko79@mail.ru

Introduction. The increase of the inflammatory skin diseases as a result of the influence of negative factors (household chemicals, hard water, the UV radiation etc.) on skin has been observed in the last few years [1, 14]. As a result of their influence there is destruction of damaged skin cells and elimination of biochemical phase alteration mediators:  serotonin, histamine, lysosomal enzymes, products of proteolysis (kinins) and lipolysis (prostoglandins, prostacyclin, etc.), the processes of lipid peroxidation (LPO) are activated, etc [2, 6, 7]. The accumulation of mediators in the skin leads to the development of secondary alteration, violation of integrity of other cells and vascular responses [2]. These biochemical changes contribute to formation of a number of clinical symptoms: congestion, edema, flaking, skin ulceration [2, 7]. Considering the peculiarities of skin inflammation and its clinical symptoms, topical therapy should be directed not only on liquidation of inflammatory symptoms, but also on recovery of the structure of the damaged skin and improvement of its barrier function [1, 13].

Topical glucocorticosteroids (GCS) are the basic medications in therapy of skin inflammatory diseases, which are powerfully and quickly suppress inflammation at all stages of its development [1, 6]. Fluorinated GCS are the most effective at the moment. However, they can cause systemic (immunosuppression, steroid diabetes, atrophy of the adrenal glands and others) and local (skin atrophy, trophic disorders, regeneration and others) side effects [1, 5, 10, 11]. In this  connection it is important to create drugs that have powerful local anti-inflammatory effect, but have low risk of side effects (SE). Nonfluorinated GCS (mometasone furoate that metilprednіzolon atseponat) is one of this medications.

The search of opportunities to optimize the containts of the given medications by the base modification (which reduces the falling GCS to blood flow) and adding to their containts the components of reparative and moisturizing effects that would restore the structure and barrier properties of the skin (for example, ceramides that act in the superficial layers of the epidermis and help to restore its lipid balance, improving the barrier function of the skin and remove its dryness and flaking)  is active now [9, 12].

Thereby in State Scientific Center of Drugs (SSCD) led by prof.         M.O. Lyapunov cream (mometasone furoate and ceramides) and ointment (methylprednisolone atseponat with ceramides) have been developed to improve the efficiency and safety of treating inflammatory skin. The aim of this work was to study the therapeutic effects of local medicines with GCS and ceramides in alternative inflammation.

The materials and methods. The object of the study were cream "Mometasone with ceramides" and ointment "Methylprednisolone with ceramides". The comparison medicines were cream "Elokom" (contains mometasone furoate, the manufacturer is "Schering-Plough", Belgium) and ointment "Advantan" (contains methylprednisolone atseponat, manufacturer is "Shering AG", Germany) [5].

Antialterative effect of the developed medications was studied on the model of aseptic inflammation of the rats skin and subcutaneous rats tissue,which can trace the effectiveness of treatment in alterative inflammation. The experiments were conducted on 36 rats with weight 200-220g. The alterative inflammation phase was caused by the subcutaneous injection of 0,5ml 9% acetic acid solution together with the intraperitoneal injection of 6% dextran solution in a dose of 300 mg/kg per animal [3, 7]. The animals were divided into 6 groups (6 rats per group): the first group – the intact animals; the second group – the control pathology; the third group – the animals, which were treated with the cream “Mometasone with ceramides”; the fourth group – the animals, which were treated with lotion “Elocom”; the fifth group – the animals, which were treated with ointment “Methylprednisolone with ceramides”; the sixth group – the animals, which were treated with ointment “Advantan”. The medicines were applied in preventive and therapeutic regimen for 3 days prior to the acetic acid  injection and after (until formation the maximal area of sores on the 8^th day). The indicator of antialternative effect was sore area (S, mm²). The effectiveness of the medications was assessed for their ability to inhibit the area of ​​ damage on the skin in comparison with the group of control pathology. The area of ​​necrosis was measured in all groups on the 8^th day of the experiment (the peak of pathology). The antialternative effect of the medications (%) was calculated by the formula: А=(S_к -S_o)/S_к×100%, where S_o – it is sore area in the group of the experiment, mm²; S_к –  in the control group, mm².

To assess the influence of medications on the alteration processes on the biochemical level at the peak of pathology the identification of protein metabolism indicators the level of total protein (TP) and creatinine – was conducted the sets of reagents of company "Filisist-Diagnosis" were used; LPO activity – in terms of TBA-reactant (a set of the firm "TBA-AGAT"); antioxidant system state (AOS) – by the level of reduced glutathione (RG) (free SH-groups by the reaction with Ellman reagent) [4]. 

The statistic analysis of the results were calculated using the software package "Statistica6" at the significance level р≤0,05 [8].

Results and discussions. The results of the study of antialterative medications activity are shown in Table 1.

The observations of the animals showed that on the 8th day after playing alternative inflammation in the groups of animals different defects by size and forms developed. Thus, in the group of animals with control pathology the most average area of sores 247,17 mm² was observed (Tabl. 1). 

In the groups of animals which were treated with cream “Mometasone with ceramides” the area of opening the sores was authentically lower at 3.7 times; with cream “Elocom” – in 3.4 times; with ointment “Methylprednisolone with ceramides” – in 4.3 times and with ointment “Advantan” – in 2,7 times in comparison with the control pathology respectively. The comparison of the medicines efficacy showed that while applying the cream “Mometasone with ceramides” the area of sores was 1.1 times lower in comparison with cream “Elocom”, and the applying the ointment “Methylprednisolone with ceramides” area of sores was 1.6 times lower in comparison with the ointment “Advantan”. Therefore, the developed medications with GCS and ceramides can suppress the development of the alterative inflammation and their antialternative effect was at 74,4%-76,7% level. On of the positive indexes of the created creams and ointment is that adding ceramides to GCS didn’t reduce their alternative effect.

Table 1

Efficiency of studying medications on the model of alterative skin inflammation of rats (n=6)

Notes:

1. ^# - reliability of index for the angular Fischer transformation relatively the CP;

2. **- reliability relatively to the CP (р≤0,05, Mann-Whitney criterion);

3. n –  number of animals in the  group;

4. MF – mometasone furoate, MPA – methylprednisolone atseponat, C – ceramides; CP – control pathology.

The development of alternative inflammation was also accompanied by changes in the biochemical parameters (Tabl. 2), the analysis of which showed that level of GP decreased authentically (1.4 times) in rats blood after sore formation in animals group with the control pathology and the creatinine level increased (in 2.0 times) relatively to intact animals. These changes indicate the level of destruction of tissue protein components, confirming the advantage of the catabolic processes over the anabolic [4]. The authentic increasing of lipid peroxidation parameters (TBA-reactant in 2,0 times) and depletion of AOC (RG level decreased in 1,5 times) indicate the intensity of inflammation.

We observed a normalization of biochemical indicators in animals groups which were treated with medications. That confirms the reduction of aseptic inflammation and process of sores formation on the animals skin. Thus, the level of GP increased authentically in applying the cream "Mometasone with ceramides" and ointment "Methylprednisolone with ceramides" – in 1.1 times and in 1.2 times respectively cream “Elocom” and ointment “Advantan” – in 1.4 times relatively to the control pathology. Also we observed a significant decrease of the creatinine level in the treatment with ointment “Methylprednisolone with ceramides” and cream "Mometasone with ceramides" in 1.4 times, with cream “Elocom” in 1.7 times and ointment “Advantan” in 1.8 times in comparison with the control pathology. The increase of GP level and reduced of the creatinine level indicate the of the anabolic processes benefits raising over the catabolic ones.

 Table 2

Biochemical parameters of rats blood with skin aseptic inflammation, (n=6)

Notes:

1.* – reliability relatively to the intact animals (р≤0,05, Mann-Whitney criterion);

2.** – reliability relatively to the control pathology (р≤0,05);

3. n – the number of animals in group;

4. MF – Mometasone furoate, MPA – methylprednisolone atseponat, C – ceramides, CP –  control pathology.

           Normalization of LPO and of AOC state (Table 2) indicates the reduce of the pathological process. Thus, the level of TBA-reactant decreased authentically under the influence of creams "Mometasone with ceramides", “Elocom” and ointment “Advantan”  in 1.5 times, and under the influence of ointment "Methylprednisolone with ceramides" in 1.6 times relatively to the control pathology. The RG level authentically increased in 1.2 times when applying cream "Elokom" and ointment "Advantan" – in 1.3 times, relatively to the controlled pathology applying cream "Mometasone with ceramides" and ointment "Methylprednisolone with ceramides".

The analysis of the biochemical studies showed that the better  normalization of the system parameters was observed when applying the cream "Elokom" and ointment "Advantan". This fact indicates that GCS can get into the systemic circulation and increase the risk of systemic adverse reactions. Normalization of biochemical parameters was observed using medications with GCS and ceramides, but they didn’t get intact values. This shows the little influence of topical medications on blood system parameters of rats during the development of pathology. Thus, by injecting ceramides that work in the surface layers of the skin and able to concentrate the active substance in the epidermis, revenues to the ACS circulation and the risk of SE decreased. In this way, it can be concluded that cream "Mometasone with ceramides" and ointment "Methylprednisolone with ceramides" can reduce sores formation on the skin (antialterative effect was 74.4% and 76.7%, respectively) and promote  the normalization of the inflammatory markers in the blood. The medications with GCS exceeded the comparison medications the cream "Elokom" and ointment "Advantan" according to the expressiveness of local effect, and the developed drug inferior the comparison medications by the systemic action, that testifies the decrease of getting GCS to the systemic circulation and reduce their toxicity and improving the safety of treatment.  

Conclusions:

1. On the model of aseptic inflammation of the skin rats it has been found that the cream "mometasone with Ceramides" and ointment "Methylprednisolone with Ceramides" had antyalteration action at 74.4% and 76.7%, respectively, and their local therapy was higher than the efficiency of the reference drugs - cream "Elokom" (70.7%) and ointment "Advantan" (63.0%).
2. It was found that the cream "Mometasone with Ceramides" and ointment "Methylprednisolone with Ceramides" at least modify the system parameters of blood, which reduces the risk of the systemic adverse reactions GCS and increases the safety of the local treatment with hormonal therapy.

4. Thus, the further study is perspective of the proposed combinations of corticosteroids with ceramides to implement them in clinical dermatology.