CHARACTERISTICS OF REDOX STATUS IN MALE AND FEMALE RATS WITH EXPERIMENTAL DIABETES MELLITUS, GUERIN'S CARCINOMA AND ASSOCIATED PATHOLOGY

Taking into account cancer and diabetes comorbidity, the role of free radicals in these conditions and the dependence of pathological processes on the gender of animals, the aim of the study was to examine lipid peroxidation (LPO) intensity and the activity of the key antioxidant enzyme superoxide dismutase (SOD) in the heart, kidneys, liver, and in tumors of male and female rats with diabetes mellitus (DM), Guerin's carcinoma (GC) or both diseases. Materials and Methods. The study included 80 white nonlinear male and female rats (180–220 g). The animals were divided into 4 groups, each group included 10 animals (either male or female). At the time of GC inoculation, the blood glucose level in animals with alloxan diabetes was 25.4±1.2 mmol/L. The authors used conventional spectrophotometric methods to examine the content of malondialdehyde (MDA), diene conjugates (DC), and SOD activity. Statistical analysis was performed using Statistical analysis software Statistica 10.0. Results. The most pronounced changes in the studied parameters were found in the heart of female rats with isolated GC and associated diabetes mellitus: more than a threefold increase in MDA, a significant increase in DC against the background of an increase in SOD activity (by 5.5–6.3 times in comparison to intact animals). The amount of MDA in GC tissue depended on the tumor size: the maximum increase in both parameters was observed in male rats with GC growth and associated DM. Conclusion. The changes in the content of LPO products and SOD activity in the heart, kidneys, and liver of rats with diabetes mellitus and tumor growth depend on the type of the examined tissue and the gender of the animals. Disorders of the redox status found in the heart tissue can make a significant contribution to cardiopathology, which is often observed in diabetes mellitus and malignant tumor growth. Key words: diabetes mellitus, Guerin's carcinoma, rats, combined pathology, heart, kidneys, liver, malondialdehyde, diene conjugates, superoxide dismutase. Учитывая коморбидность рака и диабета, роль свободнорадикальных процессов при этих состояниях и зависимость течения патологических процессов от пола животных, целью исследования явилось изучение интенсивности перекисного окисления липидов (ПОЛ) и активности ключевого антиоксидантного фермента супероксиддисмутазы (СОД) в сердце, почках, печени, а также в опухоли крыс разного пола при сахарном диабете (СД), карциноме Герена (КГ) и при их сочетании. Материалы и методы. В исследование были включены 80 белых нелинейных крыс обоего пола массой 180–220 г, разделенных на 4 группы по 10 животных каждого пола. На момент перевивки КГ уровень глюкозы в крови животных с аллоксановым СД составил 25,4±1,2 ммоль/л. Содержание малонового диальдегида (МДА), диеновых конъюгатов (ДК) и активность СОД исследованы общепринятыми спектрофотометрическими методами. Статистический анализ проведен с использованием программы Statistica 10.0. Результаты. Наиболее выраженные изменения изученных показателей выявлены в сердце самок при изолированной КГ и КГ, растущей на фоне СД: более чем трехкратное увеличение МДА, значимый прирост ДК на фоне повышения активности СОД в 5,5–6,3 раза относительно интактных животных. В ткани КГ прослеживалась зависимость степени выраженности увеличения содержания МДА от размеров опухоли: максимальное увеличение обоих показателей наблюдалось у самцов при росте КГ на фоне СД. Выводы. Направленность изменения содержания продуктов ПОЛ и активности СОД в сердце, почках и печени крыс при СД и опухолевом росте зависит от типа исследованной ткани и пола животных. Обнаруженные в ткани сердца нарушения редокс-статуса могут вносить значимый вклад в развитие кардиопатологии, часто наблюдаемой при СД и злокачественном росте. Ключевые слова: сахарный диабет, карцинома Герена, крысы, сочетанная патология, сердце, почки, печень, малоновый диальдегид, диеновые конъюгаты, супероксиддисмутаза.

Effect of diabetes mellitus on levels of insulin-like growth factors and their carrier proteins in Guerin’s carcinoma and it’s perifocal tissue in rats

Purpose of the study. Diabetes mellitus (DM) is considered an independent risk factor for higher cancer incidence and death rates. The system of insulin-like growth factors and their carrier proteins (IGF and IGFBP) and hyperglycemia create favorable conditions for the proliferation and metastasis of cancer cells.Materials and methods. Outbred male and female rats were divided into groups (n = 8 each): controls - with Guerin's carcinoma; main group - Guerin's carcinoma growing in presence of DM. Experimental DM was reproduces in animals by the single intraperitoneal alloxan injection (150 mg/kg body weight). After 10 days of the carcinoma growth, levels of IGF and IGFBP in the tumor and in it's perifocal area were measured using ELISA.Results. DM in females upregulated levels of glucose both in the tumor and in perifocal tissues by 1.8 (p < 0.05) and 8.1 times, respectively, but caused opposite changes in IGF-I - it's increase by 6.3 times in the tumor and decrease by 3.2 times in the perifocal area; as a result, such tumors with small primary nodes were more "aggressive" and actively metastasized. In males, induced DM downregulated levels of glucose, IGF-II and IGFBP2 in the carcinoma by 8.4, 3.1 and 1.7 (p < 0.05) times, respectively, and increased levels of IGF-I and IGFBP2 by 1.4 and 1.3 times (p < 0.05) in the perifocal area without changing glucose levels; as a result, tumor volumes exceeded the values in the standard growth, without metastasizing into visceral organs.Conclusion. We revealed gender differences in changing levels of glucose and IGF both in the tumor and in it's perifocal tissue in rats with Guerin's carcinoma growing in presence of DM; these differences could determine different tumor growth dynamics in male and female rats.

Structural and functional state of Heren carcinoma after local fractional X-irradiation and irradiation combined with meloxicam

Background. One of the most important problems of oncology is the overcoming of therapeutic resistance of tumors, which occurs in particular due to increased levels of the enzyme cyclooxygenase 2 (COX-2). It is known that the growth of COX-2 and the product of its activity, prostaglandin-E2 in cancer, promotes such processes in the body as tumor growth, stimulation of proliferation, induction of cancer stem cells, inhibition of apoptosis, activation of angiogenesis, invasion, metastasis and development of chemoresistance. The use of COX-2 inhibitors, which are nonsteroidal anti-inflammatory drugs (NSAIDs), significantly limits these processes and improves survival and mortality in cancer patients, and in combination with chemotherapeutics eliminates the resistance they cause. Purpose – study of the structural and functional state of Guerin’s carcinoma cells after the combined use of nonsteroidal anti-inflammatory drug meloxicam and local X-irradiation in total doses of 1.0 and 10 Gy. Materials and methods. On 33 rats with inoculated Guerin’s carcinoma, the ultrastructure of tumor cells (TC) was studied using standard methods of electron microscopy 24 hours after the combined use of the meloxicam drug at a dose of 0.2 mg per 1 kg of body weight one day before the first and 2 hours before the second session fractional local X-irradiation in total doses of 1 and 10 Gy (twice daily at 0.5 and 5.0 Gy, respectively. The mitotic index (the number of cells in the state of mitosis per 100 TC,%), the apoptosis index (the number of cells in the state of apoptotic death per 100 TC,%) and the frequency of TC with small nuclei (%). Results. It was found that irradiation of Guerin’s tumor in a total dose of 10 Gy causes disturbances in the ultrastructure associated with damage to the nuclear apparatus of the TC. Pleiomorphism of the nuclei, the appearance of binucleated cells and micronuclei, a significant decrease in mitotic activity and a slight increase in the apoptosis index are observed. Stimulation of the functional activity of macrophages is also noted. Under irradiation in a total dose of 1 Gy, such effects are less pronounced or completely absent, such as, for example, the processes of phagocytosis. The frequency index of TC with small nuclei is equally reliably increased at both radiation doses. The administration of the drug meloxicam leads to a significant decrease in mitotic activity and an increase in the frequency of small cells, while the ultrastructural picture of the tumor remains almost unchanged. With the combined action of the drug and radiation in both doses, violations of the fine structure of the OC are identical to those found during irradiation. At the same time, the mitotic index in the group with the combined effect of the drug and radiation at a dose of 10 Gy is significantly lower than with only irradiation.In addition, at both doses, the frequency of small forms of PC significantly increases in comparison with the indicators of both the intact control group and the corresponding irradiation groups. Only in combination with radiation does meloxicam reliably stimulate apoptosis, while in other groups its index remains at the level of control values. The relationship was confirmed, which was constantly revealed in all experimental groups, between a decrease in the level of the mitotic index and an increase in the frequency of TC with small nuclei in Guerin’s carcinoma. An inverse correlation was found between these indicators (r = 0.80, P = 0.05). Conclusions. The combined action of the drug and irradiation significantly increases the effectiveness of both therapeutic factors due to the property of meloxicam to reliably inhibit proliferative activity and promote post-radiation development of apoptosis in tumor tissue. The presence of a correlation between the mitotic index and the frequency of cells with small nuclei in Guerin’s tumor may indicate the relationship between cell growth and division. Under the combined action of both investigated factors, changes in the tumor ultrastructure are mainly caused by irradiation. The administration of meloxicam increases the efficiency of the combined use of both therapeutic agents due to its ability to reliably inhibit proliferative activity and promote post-radiation development of apoptosis in tumor tissue. The presence of a correlation dependence between the mitotic index and the frequency of cells with small nuclei in Guerin’s tumor may indicate the relationship between the processes of cell growth and division.

Functional state of cardiomyocyte mitochondria in malignant process in presence of comorbid pathology in experiment

Purpose of the study. An analysis of indices of free radical oxidation and respiration of mitochondria of heart cells in a malignant process in presence of diabetes mellitus and chronic neurogenic pain in experimental animals.Materials and methods. The study included outbred female rats (n=32) and С57ВL/6 female mice (n=84). Experimental groups of rats were: intact group 1 (n=8), control group 1 (n=8) with diabetes mellitus (DM), comparison group 1 (n=8) with standard subcutaneous transplantation of Guerin’s carcinoma, main group 1 (n=8) with Guerin’s carcinoma transplanted after 1 week of persistent hyperglycemia. Experimental groups of mice were: intact group 2 (n=21), control group 2 (n=21) with a model of chronic neurogenic pain (CNP), comparison group 2 (n=21) with standard subcutaneous transplantation of melanoma (B16/F10), main group 2 (n=21) (CNP+B16/F10) with melanoma transplanted 3 weeks after the CNP model creation. Heart mitochondria were isolated by differential centrifugation. Levels of cytochrome C (ng/mg of protein), 8-hydroxy-2'-deoxyguanosine (8-OHdG) (ng/mg of protein), and malondialdehyde (MDA) (μmol/g of protein) were measured in mitochondrial samples by ELISA. Statistical analysis was performed using the Statistica 10.0 program.Results. DM in rats upregulated 8-OHdG by 6.3 times and MDA by 1.9 times (р=0.0000) and downregulated cytochrome C by 1.5 times (р=0.0053) in heart cell mitochondria, compared to intact values. DM+Guerin’s carcinoma in rats increased 8-OHdG by 14.0 times and MDA by 1.7 times (р=0.0000) and decreased cytochrome C by 1.5 times (р=0.0000), compared to intact values. CNP in mice did not affect the studied parameters in mitochondria of the heart. CNP+B16/F10 in mice increased 8-OHdG by 7.1 times and MDA by 1.6 times (р=0.0000) and decreased cytochrome C by 1.6 times (р=0.0008).Conclusions. Comorbidity (diabetes mellitus, chronic neurogenic pain) together with malignant pathology aggravates mitochondrial dysfunction of heart cells with destabilization of the respiratory chain mediated by free radical oxidation processes.

Suppression of tumor development by another tumor in primary immunodeficiency in experiment.

e21583 Background: Multiple primary malignant tumors (MPMTs) are characterized by the presence of several primary neoplasms in one patient. The purpose of the study was to create an experimental model of MPMTs with one dominating tumor. Methods: The study included 21 female BALB/c Nude mice. The main group included mice with simultaneous subcutaneous inoculation of tumors: Guerin's carcinoma (0.5 million tumor cells in 0.5 ml of saline solution) under the right scapula and B16/F10 melanoma (0.5 ml of suspension diluted 1:20 in saline solution) under the left scapula. Control groups included females with melanoma or carcinoma inoculated at the same dosage and volume as in the main group. Results: In a MPMT model, tumors appeared 3 times faster than in controls and demonstrated larger volumes: melanoma – by 7.5 times, carcinoma – by 2.2 times; the survival of mice with MPMTs decreased. Carcinoma in a MPMT model metastasized to melanoma and almost completely suppressed its growth. Melanoma was represented by a small “island” of tumor tissue 3-4 mm in diameter and was located just under the skin at the site of injection of melanoma cells. The light part of the same loose pasty consistency as the dark part, with a diameter of 6-7 mm, was located around the dark “center” of melanoma. The rest part of the tumor located under the left scapula looked like an elongated grayish-pink node of a dense elastic consistency - just like the tumor located under the right scapula, which was much larger in volume. The right and left tumors did not merge with each other; there was a small distance of about 2-3 mm between them. A small lesion of caseous necrosis, 6–7 mm in diameter, was recorded in the center of the right tumor node of Guerin's carcinoma; there was no necrosis in the left tumor. Smaller size, the absence of necrosis and visually more “young” carcinoma tissue on the left indicated its later appearance than that on the right, which, in combination with the remnants of melanoma fused to the left tumor and the absence of “contact” between the left tumor and the right one, indicated the metastatic nature of Guerin’s carcinoma on the left. B16/F10 melanoma did not metastasize. Conclusions: In simultaneous subcutaneous inoculation of murine B16/F10 melanoma and rat Guerin’s carcinoma to female BALB/c Nude mice, carcinoma cells metastasized to melanoma and suppressed its growth.

DIELECTROMETRIC STUDY OF RADIATION PROTECTION ABILITY OF ULTRA-DISPERSED NANODIAMONDS

Radiation protection ability of the ultra-disperse nanodiamonds (UDD) is studied based on the measurements of dielectric permittivity of red blood cells (RBC) affected by cancer. Wistar rats with Guerin's carcinoma were treated by X-ray 5.8 Gy. Some rats received UDD with food during 5 days prior to the X-ray. The groups with UDD, X-ray, and both UDD and X-ray treatments were compared to the control group. The complex dielectric permittivity of the RBC was measured by microwave dielectrometry. It was shown, tumor development leads to the increase in the dielectric permittivity and relaxation frequency. The irradiation promotes further growth of the parameters, while UDD uptake leads to insignificant changes in comparison to the control group. Therefore, UDD occur the radioprotective effect promoting repair, compensation and restoration of body tissues that is demonstrated by normalization of the dielectric parameters of RBC.

Determination of the anticancer properties of cis- and trans-diadamanthylcarboxylates of dirhenium(III)

The aim of the study. The aim of the work was to investigate in vivo anticancer activity of cis- and trans-diadamanthylcarboxylates of dirhenium(III) alone and together with cisplatin in form of nanobins.Materials and methods. Model of tumor growth, Guerin’s carcinoma; intraperitoneal administration of cisplatin, dirhenium(III) compounds in liposomes and of binary liposomes, containing both cytostatics; volumes and final weights of tumors were measured.Results. In vivo antitumor properties of two dirhenium(III) dicarboxylates with 1-adamantanecarboxylic acid moieties as ligands with cis- (I) and trans- (II) orientation of the carboxylic groups around a cluster fragment alone and together with cisplatin were presented; an attempt to understand differences in a possible mechanism of anticancer activity of the substances were undertaken. Antiradical and DNA-binding properties of I and II were the matter of consideration.Conclusions. Cis- and trans- compounds of dirhenium I and II had close antitumor activity in vivo with a little bit superiority of the cis- analog. Mechanisms of anticancer activity of I and II are different and may also include monofunctional adduct formation and subsequent interstrand cross-linking for the II substance, formation of protein-DNA cross-links, etc.

State of enzymatic and non-enzymatic links of antioxidant protection of the liver of tumor-bearing rats under the action of laser irradiation

The activity of antioxidant enzymes — superoxidedismutase (SOD), catalase and glutathionetransferase, and also the level of low molecular weight antioxidants — vitamin E and ascorbic acid in the liver subcellular fractions of rat with transplanted of Guerin’s carcinoma it was investigated. It is shown that in the liver of tumor-bearing rats in the logarithmic phase of oncogenesis increases the activity of the components of the enzymatic link of the antioxidant system (AOS) and the content of vitamin E with a simultaneous decrease of ascorbic acid. The AOS depletionis expressed by a decrease of antioxidant enzymes activity and a decrease the level of low molecular weight antioxidants in the stationary phase of oncogenesis in the liver cells of tumor-bearing rats. It was found that laser irradiation of rats in the area of tumor growth reduces its effect on liver AOS, which is manifested by increased activity of SOD, catalase and vitamin E content in the stationary phase of Guerin’s carcinoma growth in the body.

Effect of laser irradiation on xanthine oxidase activity and superoxide radical generation in rat liver cytosol fraction

The effect of tumor growth in the body and laser irradiation on the enzymatic activity of xanthine oxidase, in particular its D- and O-forms, and also the rate of generation of the superoxide radical (O2–) and the level of protein sulfhydryl groups in the liver rat cytosolic fraction has been investigated. It has been found that in the cytosolic fraction of rats with transplanted Guerin’s carcinoma decreases the enzymatic activity of the D-form of xanthine oxidase with a simultaneous increase in its O-form during the period of intensive (14 days, which corresponds to the logarithmic phase of on cogenesis) and the period of final tumor growth (21 days, which corresponds to the stationary phase of oncogenesis). The increase in the enzymatic activity of the O-form of xanthine oxidase was accompanied by an increase the rate of superoxide radical generation and a decrease in the level of protein SH-groups in the liver cytosolic fraction of tumor-bearing rats. Daily directed action of laser irradiation on the area of growth of Guerin’s carcinoma leads to less destructive changes in the liver. Thus, there is an increase in the enzymatic activity of the D-form of xanthine oxidase, a decrease the rate of superoxide radical formation and an increase the content of protein SH-groups in the cytosolic fraction of the liver of experimental animals compared with non-irradiated tumor-bearing rats.