Background. Oxidative stress plays an important role in the development of peritonitis, which leads to a decrease in antioxidant level and an increase in oxidant production. Studying ATPase activity and lipid peroxidation when using new methods of treating experimental peritonitis is of interest for clinical practice.The aim. To study the dynamics of ATPase activity and lipid peroxidation indicators in liver tissue in experimental peritonitis when using Serogard.Methods. The study was conducted on male Wistar rats aged 6 months. All animals were subjected to experimental peritonitis using the method we developed. Animals in the control group (n = 20) were intraperitoneally injected with 3 ml of physiological solution one day after modeling peritonitis; animals in the experimental group (n = 19) were injected Serogard® (AO Pharmasintez, Russia). We determined the initial values of the indicators in healthy rats (n = 7). In animal liver tissue, total and mitochondrial ATPase activity was studied, as well as the content of malondialdehyde (MDA) and diene conjugates (DC).Results. It was found that the total and mitochondrial activity of ATPases was suppressed by the day 3 of experimental peritonitis. During 14 days of the experiment, ATPase activity did not reach normal levels in any of the groups. However, animals in the experimental group had dynamics of a more steady, significant increase in the total ATPase and the mitochondrial ATPase (ATP synthase) activity with a significantly higher level than in the control group. MDA and DC in the experimental group increased by days 7–14, which indicated the activity of free radical processes and was also an indicator of the dynamics of restoration of metabolic and energy processes.Conclusion. Using Serogard in experimental peritonitis contributed to a more effective restoration of total ATPase activity and mitochondrial ATPase (ATP synthase) and on days 7–14 of the study led to an increase in the level of free radical processes activity that accompany and indicate the process of aerobic energy restoration.

In order to find out the mechanisms of pathogenesis of degenerative-dystrophic diseases of the spine, it is of particular interest to search for body parameters which are directly or indirectly interrelated with the key factors of peripheral conversion of nidus iodothyronines and constitute a system of network interactions, affecting metabolic indicators at the local and systemic level. The aim. To search for correlations of local key factors of peripheral conversion of Ligamentum flavum iodothyronines with indicators of biochemical, hematological and hormonal blood profiles of patients with stenosing processes of the spinal canal and dural sac in the lumbar spine. Materials and methods. 33 patients (15 males, 18 females) with stenosing processes of the spinal canal and dural sac in the lumbar spine were examined (mean age – 45.73 ± 1.95 years). The expression of deiodinase genes and other candidate genes was determined in Ligamentum flavum biopsies collected during surgical treatment. Biochemical, hematological and hormonal parameters were determined in peripheral blood. The resulting data array was processed in order to find correlations between the parameters of systemic and local metabolism. Results. The relationships of deiodinases with the expression of GDF5, MMP1, MMP3 and TIMP1 in Ligamentum flavum (p < 0.05) were found. Of the hormonal profile of the blood serum, the most significant indicators were thyreotropin, free triiodothyronine and thyroperoxidase antibodies. In the biochemical profile, levels of direct bilirubin, total cholesterol, HDL and LDL cholesterol and triglycerides changed along with the expression of deiodinases. Correlative relationships with the expression of deiodinases were found for the following hematological analytes of whole peripheral blood: hemoglobin, mean corpuscular hemoglobin concentration, numbers of granulocytes, lymphocytes, eosinophils, monocytes, band neutrophils, red cell distribution width and platelet crit. The data obtained indicate the involvement of peripheral conversion factors in the pathogenetic process and provide information to form a new view on the pathogenesis of degenerative-dystrophic processes in the Ligamentum flavum of patients with stenosing processes of the spinal canal and the dural sac in the lumbar spine.

Parathyroid hormone receptors PTH1R and PTH2R are poorly studied in the pathology of the vertebral column and may be involved in both ectopic ossification of the vertebral ligaments and fibrous growth of the extracellular matrix, as well as in other unknown processes occurring in the pathological focus. Aim. To study the expression of genes encoding parathyroid hormone receptors in intraoperative Ligamentum flavum samples of patients with stenosis of the spinal canal and the lumbar dural sac of the spine and to assess the correlation with the expression of other genes active in connective tissue in conditions of developing pathology.Materials and methods. Studies of the Ligamentum flavum were performed in 33 patients (17 women, 16 men) with stenotic processes of the spinal canal and dural sac of the lumbar spine. RNA was isolated from the biopsies using Qiagen reagents (Germany). Then reverse transcription was performed. cDNA was used for setting up a polymerase chain reaction (Promega Reagents, USA) using a CFX96 thermal cycler (Biorad, USA) with specially selected specific primers.Results. The expression of genes encoding parathyroid hormone receptors PTH1R and PTH2R was detected and evaluated for the first time in intraoperative Ligamentum flavum samples of patients with spinal canal and dural sac stenosis, and their relationship with the activity of genes encoding estradiol receptors (ESR1 and ESR2) and FGFR3 was revealed. For the first time, common features of PTH1R and PTH2R gene expression, as well as their differences, were revealed. It is proved that the intensity of expression of parathyroid hormone receptors in intraoperative Ligamentum flavum biopsies does not depend on age, gender, or the presence of bone and cartilage nodes according to MRI results. The interdependence of differential changes in the expression of genes of the receptor apparatus involved in the metabolism of Ligamentum flavum tissues was revealed.

Background. Ligamentum flavum is involved in the development of stenosis of the spinal canal and dural sac, but the mechanisms underlying degenerative-dystrophic changes have not yet been sufficiently studied. Aim. To study the expression of candidate genes potentially involved in connective tissue metabolism in intraoperative samples of Ligamentum flavum of patients with stenosis processes of spinal canal and duralsac on the lumbar spine. Materials and methods. During the surgical treatment, Ligamentum flavum biopsies were collected from 33 people with stenosis processes of spinal canal and dural sac on the lumbar spine (16 men, 17 women). RNA was isolated, then reverse transcription was performed. After that, real time PCR was performed (CFX96, Biorad) with the specific primers (RealTimePrimers.com). Results. The characteristics of the expression of candidate genes that are active in the Ligamentum flavum of  the examined patient population are given: NAT1 and NAT2 acetyltransferase genes; genes that determine the intensity of local metabolism (Dio1, Dio2 and Dio3); estradiol receptor genes ESR1 and ESR2; genes encoding receptors for growth factors and parathyroid hormone. All Ligamentum flavum samples showed activity of the NAT, ESR 2, FGFR1, FGFR3, NA20, PTH1R, and PTH2R genes. In the vast majority of samples (93.9–97.0 %), the following genes were active: PDGFA, ESR1, CALCR, and PDGFB. In 21.2–39.4 % of samples, “silence” of the Dio1, Dio2, Dio3, NAT2 and GDF5 genes was detected, and only 39.4 % of samples revealed NAT1 transcripts. The highest heterogeneity of transcript content was observed for Dio1, NAT2, and Dio2. The most stable expression in Ligamentum flavum tissue and a narrow range of fluctuations were characterized by genes with a high level of activity AANAT, ESR2, NAA20 and genes with an average level of activity FGFR1, FGFR3, PTH1R and PTH2R. Conclusions . Ligamentum flavum is a promising little-studied substrate for molecular genetic research. The revealed features of gene expression provide new information about pathogenesis, and the new fundamental knowledge obtained can provide a basis for developing methods for preventing the development of epidural fibrosis.

Epidural fibrosis is a dynamic pathological process that develops and progresses over time and is a polyetiological and multifactorial complication in the postoperative period during spinal surgery.Aim of the study. To study the reparative process and the formation of epidural fibrosis and its effect on the shape of the dural sac in the area of laminectomy at different periods of wound healing.Materials and methods. To assess the dynamics of the formation of epidural fibrosis and the involvement of the dura mater in this process, laminectomy was performed at the level of the LVI vertebra in 35 male Wistar rats at the age of 4–5 months. The animals were divided into 5 periods of withdrawal from the experiment on days 3, 7, 14, 21, and 28. The histological material was studied by staining with hematoxylin and eosin. To assess the deformation of the dural sac, the ratio of the diameter to the length of the canal was assessed on histological preparations in the laminectomy zone, and these measurements were carried out in a group of intact animals at the level of the LVI vertebra (n = 7). Results. During the observation process, a change in the ratio of the sizes of the dural sac has been observed. In the period from days 3 to 14, the anterior-posterior size of the dural sac progressively decreases, in parallel, the transverse size increases, which leads to a change in its shape. On follow-up, some form correction occurs. It is possible that the change in the shape of the dural sac in terms of up to 14 days is due to edema, plasma saturation of tissues, as well as the growth of connective tissue. The subsequent decrease in edema and the restructuring of the newly formed connective tissue leads to a correction of the shape of the dural sac.Conclusion. In the work, the dynamics of morphological changes in the zone of formation of epidural fibrosis during laminectomy in the experiment was assessed. The dynamics of changes in the lumen of the dural sac was established. The early involvement of the dura mater in the pathological process, its interest during the entire observation period was noted. Also, histological preparations show the involvement of the nerve fibers of the cauda equina in the pathological process from the early stages after surgery: vacuolization and destruction of nerve fibers, adhesion of fibers to each other and to the dura mater, as well as activation of endotheliocytes in the vasa nervorum.

New data have been obtained for assessing the expression of genes of metalloproteinases and their tissue inhibitors (MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, TIMP-1 and TIMP-2) in the Ligamentum flavum in patients with lumbar stenosis of spinal canal and dural sac. The features of the metabolism of the extracellular matrix (ECM) were revealed, the data obtained were compared with those for previously studied candidate genes. The search for relationships with the features of the ECM metabolic characteristics was carried out.The aim. To study the expression of genes of metalloproteinases and their tissue inhibitors in intraoperative biopsies of the Ligamentum flavum of patients with lumbar stenosis of the spinal canal and dural sac.Materials and methods. A group of 33 people (17 women, 16 men) with lumbar stenosis of the spinal canal and dural sac was studied; the average age is 45.73 ± 1.95 years. RNA was isolated from intraoperative biopsies of the Ligamentum flavum, reverse transcription was performed, and PCR using specific primers was performed.Results. In Ligamentum flavum of patients with stenosing processes of the spinal canal and dural sac, an increased activity of MMP-1 and insufficient response of TIMP-1 and TIMP-2 were found; the expression of MMP-1 increased synchronously with Dio2, and both genes decreased their activity with increasing age of the patient. In patients with Ligamentum flavum ossification, the MMR-8 gene was more actively expressed, and the synthesis of the mRNA of the MMR-9 gene decreased compared to the subgroup without ossification.

Parathyroid hormone receptors PTH1R and PTH2R are poorly studied in the pathology of the vertebral column and may be involved in both ectopic ossification of the vertebral ligaments and fibrous growth of the extracellular matrix, as well as in other unknown processes occurring in the pathological focus. Aim . To study the expression of genes encoding parathyroid hormone receptors in intraoperative Ligamentum flavum samples of patients with stenosis of the spinal canal and the lumbar dural sac of the spine and to assess the correlation with the expression of other genes active in connective tissue in conditions of developing pathology. Materials and methods. Studies of the Ligamentum flavum were performed in 33 patients (17 women, 16 men) with stenotic processes of the spinal canal and dural sac of the lumbar spine. RNA was isolated from the biopsies using Qiagen reagents (Germany). Then reverse transcription was performed. cDNA was used for setting up a polymerase chain reaction (Promega Reagents, USA) using a CFX96 thermal cycler (Biorad, USA) with specially selected specific primers. Results. The expression of genes encoding parathyroid hormone receptors PTH1R and PTH2R was detected and evaluated for the first time in intraoperative Ligamentum flavum samples of patients with spinal canal and dural sac stenosis, and their relationship with the activity of genes encoding estradiol receptors (ESR1 and ESR2) and FGFR3 was revealed. For the first time, common features of PTH1R and PTH2R gene expression, as well as their differences, were revealed. It is proved that the intensity of expression of parathyroid hormone receptors in intraoperative Ligamentum flavum biopsies does not depend on age, gender, or the presence of bone and cartilage nodes according to MRI results. The interdependence of differential changes in the expression of genes of the receptor apparatus involved in the metabolism of Ligamentum flavum tissues was  revealed.

Background. Ligamentum flavum is involved in the development of stenosis of the spinal canal and dural sac, but the mechanisms underlying degenerative-dystrophic changes have not yet been sufficiently studied. Aim. To study the expression of candidate genes potentially involved in connective tissue metabolism in intraoperative samples of Ligamentum flavum of patients with stenosis processes of spinal canal and duralsac on the lumbar spine.Materials and methods. During the surgical treatment, Ligamentum flavum biopsies were collected from 33 people with stenosis processes of spinal canal and dural sac on the lumbar spine (16 men, 17 women). RNA was isolated, then reverse transcription was performed. After that, real time PCR was performed (CFX96, Biorad) with the specific primers (RealTimePrimers.com).Results. The characteristics of the expression of candidate genes that are active in the Ligamentum flavum of the examined patient population are given: NAT1 and NAT2 acetyltransferase genes; genes that determine the intensity of local metabolism (Dio1, Dio2 and Dio3); estradiol receptor genes ESR1 and ESR2; genes encoding receptors for growth factors and parathyroid hormone. All Ligamentum flavum samples showed activity of the NAT, ESR 2, FGFR1, FGFR3, NA20, PTH1R, and PTH2R genes. In the vast majority of samples (93.9–97.0 %), the following genes were active: PDGFA, ESR1, CALCR, and PDGFB. In 21.2–39.4 % of samples, “silence” of the Dio1, Dio2, Dio3, NAT2 and GDF5 genes was detected, and only 39.4 % of samples revealed NAT1 transcripts. The highest heterogeneity of transcript content was observed for Dio1, NAT2, and Dio2. The most stable expression in Ligamentum flavum tissue and a narrow range of fluctuations were characterized by genes with a high level of activity AANAT, ESR2, NAA20 and genes with an average level of activity FGFR1, FGFR3, PTH1R and PTH2R.Conclusions. Ligamentum flavum is a promising little-studied substrate for molecular genetic research. The revealed features of gene expression provide new information about pathogenesis, and the new fundamental knowledge obtained can provide a basis for developing methods for preventing the development of epidural fibrosis.

Актуальность. В настоящее время совершенно не изучена экспрессия генов ацетилтрансфераз в структурах, участвующих в формировании патологического очага при развитии дегенеративно-дистрофических процессов в позвоночно-двигательном сегменте. Ведущая роль гипертрофии и оссификации жёлтой связки (Ligamentum flavum) в развитии и прогрессировании стенозирующих процессов позвоночного канала и дурального мешка, а также участие ацетилтрансфераз в метаболизме соединительной ткани определило цель исследования: выявить генетические особенности реакций ацетилирования у больных со стенозами позвоночного канала и дурального мешка на поясничном отделе позвоночника в зависимости от выраженности процессов оссификации Ligamentum flavum. Методы. В исследование включен 31 пациент (15 мужчин, 16 женщин, средний возраст 45,7 ± 2,0 лет) со стенозирующими процессами позвоночного канала и дурального мешка на поясничном уровне. У всех пациентов во время оперативного вмешательства были забраны образцы Ligamentum flavum, в которых затем общепринятыми методами патоморфологии определяли признаки наличия или отсутствия гипертрофии и / или оссификации, а также методом количественной ПЦР (кПЦР) определяли экспрессию генов NAT1 и NAT2. Результаты. В биоптатах Ligamentum flavum обнаружена экспрессия генов NAT1 и NAT2, что свидетельствует о ранее неизвестной роли процессов ацетилирования в метаболизме соединительной ткани. Варианты «медленного» ацетилирования достоверно чаще встречались у пациентов с выявленной оссификацией Ligamentum flavum. Обнаружены различные паттерны экспрессии генов NAT1 и NAT2, что свидетельствует о полиморфности клинических вариантов патологии. Часть выявленных вариантов экспрессии изоферментов ацетилтрансфераз характеризуют склонность к развитию оссификации Ligamentum flavum. Женщины чаще страдают от оссификации Ligamentum flavum по сравнению с мужчинами (χ2 = 39,1, р < 0,01), при этом, повторные оперативные вмешательства, как и избыточная масса тела, не увеличивали риск её оссификации. Активность экспрессии гена NAT2 можно отнести к информативным маркёрам, свидетельствующим о повышенном риске развития процессов эктопической оссификации Ligamentum flavum. Заключение. Выявленные особенности реакций ацетилирования у больных со стенозами позвоночного канала и дурального мешка могут помочь клиницистам прогнозировать развитие гипертрофии и оссификации жёлтой связки при оперативном вмешательстве на позвоночнике, а полученные новые фундаментальные знания лягут в основу разработки способов профилактики развития эпидурального фиброза на дооперационном этапе. Background. Expression of acetyltransferase genes in structures participating in formation of a pathological focus in development of degenerative spine conditions is presently unstudied. Hypertrophy and ossification of Ligamentum flavum play a leading role in the development and progression of stenosing processes in the spinal canal and dural sac whereas acetyltransferases are involved in metabolism of connective tissue. The aim of this study was to identify genetic characteristics of acetylation reactions in patients with stenosis of the lumbar spinal canal and dural sac depending on severity of ossification of Ligamentum flavum. Methods. This study included 31 patients (15 men and 16 women aged 45.7 ± 2.0 years) with stenosing processes of the spinal canal and dural sac at the lumbar level. Samples of Ligamentum flavum were taken from all patients at the time of surgical intervention. Signs of hypertrophy and/or ossification were detected in these samples with standard pathomorphological methods. Real time PCR was used to determine the expression of NAT1 and NAT2 genes. Results. Expression of NAT1 and NAT2 genes was found in biopsy samples of Ligamentum flavum. This indicated a previously unknown role of acetylation processes in connective tissue metabolism. Variants of «slow» acetylation were significantly more common in patients with documented ossification of Ligamentum flavum. Different patterns of NAT1 and NAT2 gene expression were identified, which suggested polymorphism of clinical variants of the disease. Some of the identified variants of acetyltransferase isoenzyme expression characterize a tendency towards development of Ligamentum flavum ossification. Women were more likely to have ossification of Ligamentum flavum than men (χ2 = 39.1, p < 0.01). Repeated surgery and overweight did not increase the risk for ossification of Ligamentum flavum. The activity of NAT2 gene expression can be classified as an informative marker for increased risk of ectopic ossification of Ligamentum flavum. Conclusion. The identified patterns of acetylation reactions in patients with spinal canal and dural sac stenosis may help clinicians to predict the development of hypertrophy and ossification of Ligamentum flavum during spine surgery and to find ways for prevention of epidural fibrosis at the preoperative stage.

Background. Mechanisms are known that allow partial local regulation of the thyroid status of tissues using peripheral conversion of iodothyronines, which determines the local level of metabolic activity. The molecular mechanisms of degenerative-dystrophic changes in the ligamentum flavum are not well understood; only a few studies on the expression of genes in this substrate have been published.The aim of the study was to identify key factors in the peripheral conversion of iodothyronines in ligamentum flavum in patients with stenosis of spinal canal and dural sac in the lumbar spine and to evaluate the features of the expression of deiodinase genes depending on the severity of ossification processes of ligamentum flavum.Methods. A group of patients with stenotic processes of the spinal canal was examined: 31 people (15 men, 16 women). The average age was45.73 ± 1.95years. In intraoperatively collected samples of ligamentum flavum, the expression ofdeiodinase genes (real time PCR) was studied and the severity of ossification processes was evaluated using a histopathological study.Results. The key factors of peripheral conversion of iodothyronines in intraoperative bioptates of the ligamentum flavum of patients (n = 31) with stenosis of the spinal canal at the lumbar level were revealed. Information was obtained on the local level of metabolic activity in the focus of pathology. The intensity and variability of deiodinase gene expression in ligamentum flavum samples were evaluated. The distribution of deiodinase gene expression patterns in groups formed on the basis of presence/absence of ossification of ligamentum flavum was found. Various variants of peripheral conversion of iodothyronines in biopsy samples of the yellow ligament of the operated patients were determined and the frequency of their occurrence was estimated.Conclusion. The key factors ofperipheral conversion of iodothyronines in intraoperative biopsy samples of the ligamentum flavum of patients with degenerative diseases of the spine were revealed for the first time. The reasons for the differences in tissue metabolism of iodothyronines and the place in the pathogenesis of stenotic processes are yet to be found.

In the treatment of severe osteoarthritis, endoprosthesis replacement is widely used. This method makes it is possible to select samples of synovial membrane for the research. In osteoarthritis, all components that form the joint are affected. However, if the role of cartilage tissue has been actively studied for many years, the study of the role of synovial membrane has not been given enough attention. In this regard, the aim of the study was to identify the gene expression profile by transcript content in intraoperative samples of synovial tissue of patients with stage III osteoarthritis of hip joint. During total hip replacement surgery, the fragments of synovial membrane were collected into solution that prevents hydrolysis of nucleic acids. After degradation and homogenization of the obtained biopsies, RNA was isolated using real time PCR with specific primers. The expression profile of a sample of genes involved in connective tissue metabolism in intraoperatively taken 10 samples of synovial membrane of patients with stage III osteoarthritis of hip joint was determined. The following genes are most actively expressed in the samples of synovial membrane in osteoarthritis of hip joint: FGFR1, PDGFA, NAA20, PDGFB, Dio1, ESR1, ESR2. The average expression level was registered for CALCR, Dio2, AANAT, PTH2R, NAT2. Significantly fewer transcripts in the synovial tissue were kept for the FGFR3 gene, Dio3, NAT1, PTH1R, and GDF5. The data of the correlation analysis indicates the close interconnection of metabolic pathways which include selected candidate genes. Identified tension of a network interactions indicates a decline in metabolic “flexibility” processes in the synovial tissue in terms of pathology.