RU2601919C1 - Method for correction of isoniazid-induced hepatotoxic reactions - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to medicine and aims at correction of isoniazid-induced hepatotoxic reactions. Method comprises using amaranth seed oil, produced by cold pressing of germs and shells of amaranth seeds, in a dose of 200 mg (2 gelatine capsules) 3 times a day, 1 hour before meals until clinical-laboratory values is normalized and another 1-2 weeks. At extension of the anti-tuberculosis therapy the said is used as continuous assistance with the interval between courses of 1 month.

EFFECT: method allows reducing intoxication symptoms and normalizing metabolic and morphological hepatic disorders at isoniazid-induced acute disorders.

1 cl, 3 dwg, 2 ex

Description

The invention relates to medicine and pharmacology and can be used to correct hepatotoxic reactions induced by isoniazid.

One of the main methods of complex treatment of tuberculosis patients is a course of anti-tuberculosis chemotherapy (V. Mishin. Side effect of anti-TB drugs under standard and individualized chemotherapy regimens. - M., 2004. - 208 p.), However, according to an analytical review of the main statistical indicators for Tuberculosis in the Russian Federation (2011) was recognized as effective only in 53.5% of newly diagnosed patients and in 43.9% of patients with recurrence of the disease. Among the reasons for the low efficiency of treatment of tuberculosis patients along with drug resistance, one should single out a high incidence of side effects that limit the conduct of full chemotherapy (V. Mishin. Side effect of anti-TB drugs under standard and individualized chemotherapy regimens. - M., 2004. - 208 p. .).

In many treatment regimens, to date, one of the main drugs remains isoniazid, which is the only drug used as monotherapy during primary and secondary chemoprophylaxis of tuberculosis contacts and infected persons. Along with high antibacterial activity, isoniazid has a specific organotropy and high toxicity. According to the frequency of occurrence, severity of the consequences and outcome of treatment, drug-induced (toxic) liver lesions dominate in the spectrum of complications of isoniazid chemotherapy.

For the treatment of toxic liver lesions with anti-tuberculosis drugs, a method is known consisting in the appointment of basic therapy (sparing regimen, liver diet, drinking 5% glucose solution) and the use of drugs with hepatoprotective activity, containing milk thistle (legal, carlsil, etc.) or essential phospholipids (Essential). However, published evidence of an increase in cholestasis and lack of anti-inflammatory activity (Okovity S.V. Clinical pharmacology of hepatoprotectors // Practician, 2002. - No. 3). Heptral, liv 52, silymarin, galstena were used.

A known method of reducing and preventing hepatotoxic reactions, which consists in the appointment as an additional means of the herbal preparation "Populin" (an aqueous extract of aspen bark and hodgepodge) (RU 2314113 C2, 2006).

A known method for the correction of endogenous intoxication and the prevention of undesirable side reactions of anti-TB drugs using enterosorbents (Fetisov Yu.I. Studying the mechanisms of endotoxemia in various forms of pulmonary tuberculosis before and after treatment: Ph.D. dissertation / Candidate of Medical Sciences / Yu.I. Fetisov . - Novosibirsk, 1997. - 23 pp. Possibilities of using the carbon-mineral enterosorbent SUMS-1 in pulmonology and phthisiology: Practical physician library / Kolpakov MA, Kolpakova TA, Letyagina EA, Volkhonskaya T.E. , Danilkina S.T., Bashkirova Yu.V. - Novosibirsk: Manuscript Publishing House, 2003. - 32 p.; Enterosorption / Edited by N. A. Belyakov. - L., 1991. - 336 p.).

The disadvantage of using enterosorbents is a violation of the absorption of vitamins from food, a violation of the motor function of the gastrointestinal tract, which can lead to the development of persistent constipation.

Another method is known, which consists in canceling anti-TB drugs and oral administration of ursosan at a dose of 20 mg / kg once and enterosgel at a dose of 10-15 g for 3 days, followed by a decrease in the dose of ursosan to 12-15 mg / kg per day 2-3 doses and enterosgel in age dosages against the background of the resumption of taking anti-TB drugs and are treated with ursosan to a value of alanine aminotransferase (AlAT) that does not exceed the value of the upper limit of the norm (RU 2473346, A61K 31/695, 2013).

A known method of stimulating the regeneration of hepatic tissue, which consists in the introduction of a complex of preparations containing 0.5 ml of pyridoxine hydrochloride, 2 μg of cyancobalamin, 0.5 mg of vitamin PP per 1 ml of extract of the extract of the bark of pork or fetal adrenal glands of a person (RU 2185835, A61K 35/55 , 2002).

For the prevention of hepatotoxic reactions caused by the use of drugs of the isonicotinic acid hydrazide group (GINC) (including isoniazid) and rifampicin, a method is known involving the use of the Glutoxim preparation (RU 2197984 C1, 2001).

Another way to limit complications and side effects in the treatment of tuberculosis with drugs of the isonicotinic acid hydrazide group (GINC) (including isoniazid), in which iron oxide sulfate is used in therapeutic doses to reduce intolerance to the drugs used, which are taken in advance for several at least two days before the start of anti-tuberculosis chemotherapy, then continue its administration with the GINC preparation until the end of treatment (RU 2217137 C2, 2001).

However, the effectiveness of the above methods for the treatment of toxic liver lesions is not sufficiently proven in multicenter clinical trials.

The objective of the invention is the correction of hepatotoxic reactions induced by the use of isoniazid, and, thereby, increasing the effectiveness of treatment of patients with tuberculosis.

The technical result consists in reducing symptoms of intoxication, as well as normalizing metabolic and morphological disorders of the liver in acute lesions induced by isoniazid in a shorter time.

The technical result is achieved by the fact that the method for the correction of hepatotoxic reactions induced by isoniazid involves the use of amaranth seed oil obtained by cold pressing of the amaranth seeds and shells along with the introduction of isoniazid. at a dose of 200 mg (2 gelatin capsules) 3 times a day 1 hour before meals until the normalization of clinical and laboratory parameters and another 1-2 weeks. With the continuation of anti-tuberculosis therapy, it is used as a constant accompaniment with a break between courses of 1 month.

The oil (RU 94044961, C11B 1/10, A23K 1/14, 1996) isolated by cold pressing from the seeds and shells of amaranth seeds contains phospholipids, tocopherol (vitamin E), unsaturated fatty acids, squalene, phytosterols and sterol esters and carotenoids.

Amaranth seed oil has a wide range of pharmacological activity. Known antitoxic and hepatoprotective activity of amaranth seed oil (E. Muzalevskaya. Initial assessment of antitoxic and hepatoprotective activity of amaranth seed oil. Tailed / E. Muzalevskaya, V.A. Nikolaevsky // Problems of developing new drugs: materials of the First All-Russian Scientific and Practical conferences of young scientists (June 3-5, 2013). - Moscow, 2013. - P. 74), lipid-lowering and hypocholesterolemic effect (Korean I.M. Amaranth oil reduces the total cholesterol in the blood / I.M. Koreansk Aaya, A.P. Salei, AM Makeev // Ways and Forms of Improving Pharmaceutical Education: Materials of the Interregional Scientific and Methodical Conference “Pharmaceutical Education 2003” (April 21-23, 2003). - Voronezh, 2003. - С 178-180 .; Korean I.M. Comparative pharmacological analysis of the effect of amaranth and linseed oil on the dynamics of rat lipid metabolism under cholesterol loading / I.M. Koreyanskaya, V.Yu. Sulin, EF Safonova, A.N. Postka // Bulletin of Voronezh State University: a series of chemistry, biology, and pharmacy. - No. 1. - 2006. - P. 204-208), antioxidant effect (G. N. Bliznetsova. The role of free radical oxidation processes in the mechanism of hepatoprotective action of amaranth seed oil / G. N. Bliznetsova, M. I. Retzky, N. D. Polyakova-Semenova , I.M. Koreanskaya // Biomedicine. - 2006. - No. 2. - P. 105-112), antibacterial activity (Korean I.M., Study of the antibacterial activity of amaranth oil / I.M. Koreanskaya, I.E. Izmalkova // Health problems of schoolchildren and students.New scientific trends in medicine and pharmacy: Proceedings of the International Conference (February 6-7, 2008) - Voronezh, 2008, - P. 2 26-227), anti-burn action (Korean I.M. Amaranth oil as an anti-burn agent / I.M. Koreanskaya, T.A. Gorokhova, S.N. Solennikova et al. // Actualni vymozenosti vedy - 2012, Materialy VIII Mexinorodni vedecko-practiKa conference. 06/27/2012 - 07/05/2012 Dil. 17. Biologicke vedy Chemie a chemicka technologiei. - Praha: Publishing House "Education and Science" sro 2012 - S. 5-6), cardioprotective effect (Yelisyeyeva OP, Semen KO, Ostrovska GV, Kaminskyy DV, Sirota TV, Zarkovic N, Mazur D, Lutsyk OD, Rybalchenko K, Bast A. / The effect of Amaranth oil on monolayers of artificial lipids and hepatocyte plasma membranes with adrenalin-induced stress. // Food Chemistry, Vol. 147, No. 15, 2014, p. 152-159). A tool is proposed for the prevention and treatment of chronic liver diseases (RU 2526172, 2014).

A tool is proposed for the prevention and treatment of chronic liver diseases (RU 2526172, 2014).

However, we have not found a source that describes the use of amaranth seed oil for the correction of hepatotoxic reactions induced by isoniazid.

The proposed method was studied in an experiment on white non-linear rats of males, sexually mature of productive age (3 months), kept under standard vivarium conditions in natural light, synchronized in nutrition with free access to water. Work with animals is carried out in accordance with the Order of the Ministry of Health of the Russian Federation No. 267 dated 06/19/2003 "On approval of the rules of laboratory practice."

Modeling of intoxication was carried out by intragastric administration to animals after food deprivation of isoniazid in the form of 1% starch mucus at a dose of 542 mg / kg body weight once a day for 6 days (control group). Amaranth seed oil was administered for therapeutic purposes 24 hours after the last injection of isoniazid at a dose of 50 mg / kg once a day for 3 days (experimental group). The reference hepatoprotector containing essential soybean phospholipids (EFL) was used as a comparison drug (experimental group) - Essentiale N preparation (Sanofi-Aventis S.A., Spain), administered as an official solution intragastrically at a dose of 80 mg / kg a similar scheme (Guidelines for preclinical studies of drugs. Part one. Edited by A.N. Mironov. - M.: Grif and K, 2012. - 944 p.). The introduction of these substances was carried out using a metal atraumatic gastric tube.

Statistical processing of the obtained data was carried out using the Statistica 5.0 licensed application software package on a Pentium Dual Core personal computer. Using the methods of mathematical statistics generally accepted in biology and medicine, the average values of the variational series and the error of the mean were determined. The significance of differences between the corresponding indices in the intact, control, and experimental groups was evaluated using Student's t-test.

In FIG. Table 1 is presented in Table 1, including data on the effect of the introduction of amaranth seed oil on changes in the level of feed and water consumption in rats with acute toxic liver damage induced by isoniazid (EFL - a preparation of essential phospholipids; AM - amaranth seed oil; ^* - p <0.05; ^** - p <0.01 - significance of differences when compared with the original; ⁺ - p <0.05; ⁺⁺ - p <0.01 - significance of differences when compared with control; ^• - p <0.05 - significance of differences when compared with a comparison drug (EFL); in Fig. 2 - Table 2 containing data on the change in biochemical pok of blood serum, prothrombin time and liver weight in acute toxic liver damage induced by isoniazid (EFL - a preparation of essential phospholipids; AM - amaranth seed oil; ^* - p <0.05; ^** - p <0.01 - significance of differences when comparing with intact; ⁺ - p <0.05; ⁺⁺ - p <0.01 - significance of differences when comparing with control; ^• • - p <0.01 - significance of differences when comparing with comparison drug (EFL)); in FIG. 3 - histological picture of the liver of rats with the introduction of amaranth seed oil against the background of acute toxic damage to the liver induced by isoniazid, color - hematoxylin-eosin, uv. × 100 (A - control; B - preparation of essential phospholipids, C - amaranth seed oil).

The invention is illustrated by the following example.

Example 1. The study of the hepatoprotective activity of amaranth seed oil on the background of acute toxic liver damage induced by isoniazid.

The study was performed on 60 white outbred sexually mature male rats weighing 205.0 ± 9.2 g, which were randomly divided into 10 groups of 6 in each: 6 control groups, 2 experimental groups and 2 comparison drug groups.

The criteria for evaluating the effectiveness were: dynamics of feed and water consumption, indicators of blood biochemical analysis: markers of hepatic cytolysis - alanine aminotransferase (AlAT), aspartate aminotransferase (AsAT); blood coagulation (prothrombin time), pathological, morphological and histochemical examination of the liver.

The activity of AlAT and AsAT in blood serum (E / L) was determined on a Furuno 270 open-type biochemical analyzer using the kinetic method (UV test according to IFCC method) using the Fluitest GPT diagnostic kits.

The determination of prothrombin time was carried out using the Diagem-P diagnostic kit.

We calculated the weight coefficient of the liver (the ratio of the mass of the organ to the mass of the animal). For histological studies, liver fragments were fixed in a 10% solution of neutral formalin, after which, according to the generally accepted method, they were poured into paraffin, serial sections 3-4 microns thick were prepared from paraffin blocks. The preparations for studying the general morphological structure were stained with hematoxylin and eosin.

For histochemical studies, liver fragments were fixed in 10% formalin solution with the addition of calcium chloride, after which serial sections 3-4 microns thick were prepared using a freezing microtome according to the generally accepted technique. Staining of drugs for the detection of lipids was carried out with the dye Red O.

Analysis of histological and histochemical preparations was carried out using a biological light microscope Levenhuk (Russia). The shooting was carried out using a fixed USB camera for the Levenhuk 8 Mpixels microscope connected to a personal computer.

The clinical picture of intoxication in the control group of animals was characterized by physical inactivity, lethargy, ruffled hair, a pronounced decrease in feed intake and an increase in water consumption.

Six-fold administration of isoniazid starting from the 7th day of observation was accompanied by a decrease in the proportion of feed and water consumed by 91.0% (p <0.01) and 65.0% (p <0.01), respectively, with the same values on the 10th day of the study. By 14 days, the proportion of feed consumed was 60.4% of the original, but this indicator was significantly lower than the intact index (Fig. 1).

In the experimental groups of animals treated with Essentiale N and amaranth seed oil, the animals were active by 10 days of observation, the proportion of feed consumed corresponds to the level of healthy animals while maintaining performance for 14 days against the background of increased water consumption (Fig. 1).

Thus, the introduction of amaranth seed oil for therapeutic purposes improves the overall clinical condition of animals and reduces the severity of intoxication symptoms observed with the introduction of high doses of isoniazid (acute toxic liver damage).

When conducting biochemical studies of blood serum in the control group of animals, it was found that six-time administration of isoniazid on the 10th day of observation was accompanied by a decrease in the activity of AlAT and AsAT, respectively, by 66.7% and 50.5% (p <0.05). The concentration of total bilirubin was significantly lower than intact by 33.6% (p <0.01) (Fig. 2). At the same time, diffuse fatty degeneration of the liver was histochemically detected. When conducting pathological and morphological studies, it was found that on the 10th day of the study, the animal’s stomach was enlarged and filled with undigested food masses, the liver mass was increased by 21.7% (p <0.01) relative to the intact index. This was accompanied by morphological disorders of the lobar-beam structure of the liver, the development of protein and fatty degeneration and necrobiosis of hepatocytes in most of the hepatic lobule (Fig. 3A), which generally indicates the development of experimental acute drug damage to the liver (toxic hepatitis) with isoniazid.

On the 14th day of the study, an increase in AlAT activity was observed in blood serum by 132.4% (p <0.05) relative to 10 days of the study, however, its level was lower than intact by 22.6% (p <0.05). Moreover, the activity of AsAT exceeded intact by 39.64% (p <0.01). Prothrombin time was higher than intact by 57.5% (p <0.05) (Fig. 2). Liver mass is significantly lower than intact by 13.0% (p <0.01), which indicates the development of destructive processes in the liver parenchyma. During histological examination of liver preparations, the beam structure was weakly expressed; residual effects of fatty degeneration were observed on sections stained with Red O.

The introduction of Essential H to animals with toxic hepatitis induced by isoniazid on the 10th day of observation was accompanied by a normalization of the activity of AcAT, and in some cases, an increase in the activity of AlAT by 35.9% (p <0.05) relative to the control. The content of total bilirubin in blood serum relative to intact and control increased 2.9 times (p <0.05) and 4.4 times (p <0.01), respectively (Fig. 2). The revealed changes in liver mass were statistically insignificant. On liver preparations, the beam structure of the lobules was within normal limits, the liver was full-blooded, no signs of necrobiosis were detected (Fig. 3B). On sections stained with Red O - non-generalized fatty degeneration.

On the 14th day of the study, despite pronounced hyperbilirubinemia (the concentration of total bilirubin was significantly higher than intact and control by 69.1% (p <0.01) and 37.9% (p <0.05), respectively), differences in comparison with intact activity level aminotransferases were not significant (Fig. 2). The prothrombin time was significantly lower than the control by 13.1% o (p <0.05). In the histological picture of the liver around the bile ducts, the presence of lymphoid infiltrates was noted.

The 3-day use of amaranth seed oil in animals with toxic hepatitis caused by isoniazid had a positive effect on enzymes-markers of liver damage, contributing to the normalization of AlAt and AsAt activity on the 10th day of the study (Fig. 2). The level of total bilirubin in the blood serum, in contrast to the control group and the comparison drug, corresponded to the indices of the animals of the intact group. The revealed changes in liver mass were statistically insignificant, the stomach was of a normal size. Normalization of biochemical parameters was accompanied by restoration of the histoarchitectonics of the organ. Medium-sized hepatocytes with a well-structured round nucleus and stained cytoplasm (Fig. 3B). On sections stained with Red O - residual effects of fatty degeneration. In the future, that is, on the 14th day of observation, the biochemical indicators listed above did not practically differ from their value observed by us on the 10th day (Fig. 2). In addition, on the 14th day of the study, the value of prothrombin time was significantly lower than the control by 12.8% (p <0.05). The above indicates that the introduction of amaranth seed oil in acute toxic liver damage with isoniazid significantly reduces the liver regeneration period by 28.5% (p <0.05), superior in effectiveness to the preparation of essential phospholipids (Essential H).

Thus, the proposed method can effectively reduce the severity of metabolic and morphological disorders of the liver in acute liver damage induced by isoniazid.

Example 2

In order to stabilize, increase the shelf life, the accuracy of dosing and correct unpleasant taste, it is proposed to use amaranth seed press oil in the form of a dosage form - white opaque hard gelatin capsules containing 1.0 ml amaranth seed oil; capsule size No. 000.

Claims (2)

1. A method for the correction of hepatotoxic reactions induced by isoniazid includes, along with the administration of isoniazid, the use of amaranth seed oil obtained by cold pressing of the seeds and shells of amaranth seeds at a dose of 200 mg 3 times a day 1 hour before meals until normalization of clinical and laboratory parameters and another 1-2 weeks. 2. The method according to p. 1, characterized in that when continuing anti-tuberculosis therapy, amaranth seed oil is used as a constant accompaniment with a break between courses of 1 month.

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