Enhanced CD8+ T-cell response in mice immunized with NS1-truncated influenza virus

Influenza viruses with truncated NS1 protein stimulate a more intensive innate immune response compared to their wild type counterparts. Here, we investigate how the shortening of the NS1 protein influence the immunogenicity of the conserved T-cellular epitopes of influenza virus. Using flow cytometry, we showed that the intraperitoneal immunization of mice with influenza virus encoding 124 N-terminal amino acid residues of the NS1 protein (A/PR8/NS124) induced higher levels of CD8+ T-cells recognizing immunodominant (NP366-374) and sub-immunodominant (NP161-175, NP196-210, HA323-337, HA474-483, NA427-433) epitopes compared to immunization with the virus expressing full-length NS1 (A/PR8/full NS). It is noteworthy that the response to the immunodominant influenza epitope NP366-374 was achieved with the lower immunization dose of A/PR8/NS124 virus compared to the reference wild type strain. Despite the fact that polyfunctional CD8+ effector memory T-lymphocytes simultaneously producing two (IFNγ and TNFα) or three (IFNγ, IL2, and TNFα) cytokines prevailed in the immune response to both viruses, the relative number of such T-cells was higher in A/PR8/NS124-immunized mice. Furthermore, we have found that polyfunctional populations of lymphocytes generated upon the immunization of mice with the mutant virus demonstrated an increased capacity to produce IFNγ compared to the corresponding populations derived from the A/PR8/full NS-immunized mice. Therefore, immunization with the attenuated influenza virus encoding truncated NS1 protein ensures a more potent CD8+ T-cell immune response.Influenza viruses with truncated NS1 protein stimulate a more intensive innate immune response compared to their wild type counterparts. Here, we investigate how the shortening of the NS1 protein influence the immunogenicity of the conserved T-cellular epitopes of influenza virus. Using flow cytometry, we showed that the intraperitoneal immunization of mice with influenza virus encoding 124 N-terminal amino acid residues of the NS1 protein (A/PR8/NS124) induced higher levels of CD8+ T-cells recognizing immunodominant (NP366-374) and sub-immunodominant (NP161-175, NP196-210, HA323-337, HA474-483, NA427-433) epitopes compared to immunization with the virus expressing full-length NS1 (A/PR8/full NS). It is noteworthy that the response to the immunodominant influenza epitope NP366-374 was achieved with the lower immunization dose of A/PR8/NS124 virus compared to the reference wild type strain. Despite the fact that polyfunctional CD8+ effector memory T-lymphocytes simultaneously producing two (IFNγ and TNFα) or three (IFNγ, IL2, and TNFα) cytokines prevailed in the immune response to both viruses, the relative number of such T-cells was higher in A/PR8/NS124-immunized mice. Furthermore, we have found that polyfunctional populations of lymphocytes generated upon the immunization of mice with the mutant virus demonstrated an increased capacity to produce IFNγ compared to the corresponding populations derived from the A/PR8/full NS-immunized mice. Therefore, immunization with the attenuated influenza virus encoding truncated NS1 protein ensures a more potent CD8+ T-cell immune response

Prospects for prevention of adhesion process during cardiac surgical interventions

The article is devoted to the problem of prevention of adhesions in cardiac surgery. It was determined that the problem is urgent due to the increase in the number of heart surgeries. The formation of adhesions is a reaction of the body after surgery, which is a stage of healing and partly performs a protective function. Nevertheless, the presence of adhesions violates the mechanical properties of the heart, negatively affects central hemodynamics, complicates the surgeon’s task during repeated surgical interventions and increases the risk of repeated operations.It has been shown that at present, for the prevention of adhesions, researchers tend to use biodegradable barrier materials with biocompatibility and the ability to dissolve after performing the barrier function. The main anti-adhesion agents used in cardiac surgery are membranes and gels. The requirements for an “ideal” agent for the prevention of adhesion were determined: biocompatibility, no irritating effect, no effect on wound healing, suppression of the growth of connective tissue in the pericardium.Conclusions. Until now, none of the funds has all the necessary qualities to prevent adhesion in the pericardium. Therefore, the search for effective methods for the prevention of postoperative adhesions remains relevant for cardiac surgery

Enhancement of the immunogenicity of influenza A virus by the inhibition of immunosuppressive function of NS1 protein

The truncation of the nonstructural NS1 protein is a novel approach for the generation of immunogenic attenuated influenza viruses. However, the innate immune mechanisms that cause the increased immunogenicity of influenza viruses with altered NS1 proteins are poorly understood. The goal of this study was to compare the immune responses in mice immunized with two variants of the influenza A/Puerto Rico/8/1934 (A/PR8) virus: the wild type virus (А/PR8/full NS) and the variant with the NS1 protein shortened to 124 amino acid residues (А/PR8/NS124). The investigated parameters of immunity included cytokine production, the dynamic variation of the innate immune cell populations, and the rate of the influenza-specific T-cell responses. An intraperitoneal route of immunization was chosen due to the variability in the replication capacity of the investigated viruses in the respiratory tract. The levels of interferon β (IFNβ), tumor necrosis factor α (TNFα), monocyte chemo-attractant protein 1 (MCP1), interleukin 6 (IL6), and IL27 in peritoneal washings of mice immunized with А/PR8/NS124 were significantly higher compared to the mice immunized with the wild-type virus. The А/PR8/NS124 treated group showed a delayed attraction of monocytes and neutrophils as well as a more pronounced reduction in the percentage of dendritic cells in the peritoneal cavity. The expression level of the CD86 activation marker on the cells expressing the molecules of the major histocompatibility complex II (MHCII+) was significantly higher in mice immunized with А/PR8/NS124 than in the group immunized with А/PR8/full NS. Finally, immunization with А/PR8/NS124 led to an increased formation of influenza-specific CD8+ effector T-cells characterized by the simultaneous production of IFNγ, IL2, and TNFα. We hypothesize that elevated cytokine production, enhanced dendritic cell migration, and increased CD86 expression on antigen-presenting cells upon immunization with А/PR8/NS124 lead to a more effective presentation of viral antigens and, therefore, promote an increased antigen-specific CD8+ immune response.The truncation of the nonstructural NS1 protein is a novel approach for the generation of immunogenic attenuated influenza viruses. However, the innate immune mechanisms that cause the increased immunogenicity of influenza viruses with altered NS1 proteins are poorly understood. The goal of this study was to compare the immune responses in mice immunized with two variants of the influenza A/Puerto Rico/8/1934 (A/PR8) virus: the wild type virus (А/PR8/full NS) and the variant with the NS1 protein shortened to 124 amino acid residues (А/PR8/NS124). The investigated parameters of immunity included cytokine production, the dynamic variation of the innate immune cell populations, and the rate of the influenza-specific T-cell responses. An intraperitoneal route of immunization was chosen due to the variability in the replication capacity of the investigated viruses in the respiratory tract. The levels of interferon β (IFNβ), tumor necrosis factor α (TNFα), monocyte chemo-attractant protein 1 (MCP1), interleukin 6 (IL6), and IL27 in peritoneal washings of mice immunized with А/PR8/NS124 were significantly higher compared to the mice immunized with the wild-type virus. The А/PR8/NS124 treated group showed a delayed attraction of monocytes and neutrophils as well as a more pronounced reduction in the percentage of dendritic cells in the peritoneal cavity. The expression level of the CD86 activation marker on the cells expressing the molecules of the major histocompatibility complex II (MHCII+) was significantly higher in mice immunized with А/PR8/NS124 than in the group immunized with А/PR8/full NS. Finally, immunization with А/PR8/NS124 led to an increased formation of influenza-specific CD8+ effector T-cells characterized by the simultaneous production of IFNγ, IL2, and TNFα. We hypothesize that elevated cytokine production, enhanced dendritic cell migration, and increased CD86 expression on antigen-presenting cells upon immunization with А/PR8/NS124 lead to a more effective presentation of viral antigens and, therefore, promote an increased antigen-specific CD8+ immune response

EFFECT OF JNK MAPK ON THE REPAIR OF DAMAGED SKELETAL MUSCLE

Regeneration of muscles after injuries, as well as the development of methods that stimulate this process, is an important problem in medicine and biology. The objective of the study was to evaluate the effect of local blocking of mitogen-activated protein kinase (MAPK) activity of the JNK group (c-Jun N-terminal kinase) on the repair of muscle tissue. Materials and methods. The effect of the JNK MAPK SP600125 blocker on the repair of muscle tissue was studied on a model of a skin and muscle wound in Wistar rats. The main group (n = 30) was injected with a drug plate containing SP600125 with a slow release of the active substance, the control group (n = 30) – the plate without the active substance. The number of dividing myosatellites and muscle kidneys in the damage zone was estimated. Results. Experimental studies have shown that when using a drug plate containing a JNK SP600125 blocker with a slow release of the active substance, the number of dividing myosatellites and forming muscle kidneys in the injury zone of the muscle in the main group on the 7th, 14th and 30th days was significantly higher (p < 0.05) than in the control. Conclusion. Local blockade of JNK MAPK in the zone of muscle damage provides the ability to stimulate the repair of damaged skeletal muscle

Dynamics of the Activity of MAP-Kinase Cascades in the Healing Process of Postoperative Musculocutaneous Wounds

Background. Management of the reparative process is an urgent task of modern medicine. In our opinion, the development of pathogenetically grounded approaches to optimizing the repair process for managing the interrelations of stromal cells is promising. One of the promising areas in this regard is the impact on the MAPK-cascades.Aim: to study the expression of MAP-kinase mechanisms in the regulation of repair by the example of a musculocutaneous wound.Methods. A linear muscular skin wound was modeled using Wistar rats weighing 220–250 g at the age of 9 months (n = 24). Immunofluorescence staining was performed to detect the activity of p38, JNK, and ERK MAPK cascades from 1 to 30 days.Results. It was established that specific staining in the area of connective tissue formation during staining with p38 MAPK and its phosphorylated form was first observed on the 3rd day, and its maximum severity occurred at the same time. On the 7th and 14th day, small zones in the area of scar formation were minimally stained. The phosphorylated part of the JNK-cascade in the zone of traumatic injury was detected starting from the 1st day after the injury. Bright color persisted on the 3rd day. On the 7th day, the color was minimal, and by the 14th day a second wave of expression was observed. ERK-staining was observed from the 1st to the 14th day with a peak activity on the 3rd day.Conclusion. Thus, we revealed the simultaneous involvement of p38, JNK-, and ERK-cascades in the regulation of the reparative process in the conditions of a musculoskeletal wound. At the same time, it is noteworthy that the peak activity of all cascades coincides and falls on the 3rd day

Postconditioning as a method tissue survivability enhancement in ischemic damage

The article presents the present-day data on the phenomenon of ischemic myocardial postconditioning. It has been shown, that this phenomenon consists in protection of the heart from reperfusion damage by the means of short episodes of ischemia/reperfusion performed at the early stage after prolonged ischemia. It is presented that postconditioning effect manifests in limiting the size of infarction and preserving endothelial function in the region exposed to ischemic injury. The article reports on the modern concept of role of various intracellular signal cascades in providing survival of the cell after the episode of ischemia/reperfusion. Much attention is given to changing in the state of pores localized in internal mitochondrial membrane as end links of realization of postconditioning effect. Prospects of clinical use of postconditioning are very optimistic due to the fact that application of different variants of preconditioning is limited because in most cases it is impossible to predict the time of occurrence of ischemic injury, whereas postconditioning may be used after prolonged ischemia

Angiogenesis as an adaptive mechanism in ischemia

The article is devoted to angiogenesis as one of the ways to increase the stability of the ischemic tissue. It is shown that the reduction in ischemic tissue damage is based on four basic processes - vasculogenesis of endothelial progenitor cells, angiogenesis and arteriogenesis with the growth and subsequent stabilization of the outgrowths of mural cells and the growth of collaterals (expansive growth of pre-existing vessels). However, despite the great progress in the understanding of ischemia-induced angiogenesis, and a lot of positive results from its stimulation using the disclosed arrangements natural response to the reduction of tissue perfusion is still an unsolved problem is the practical use of this knowledge to improve the metabolic processes of organs and tissues with compromised vascular bed. The effect of a variety of recombinant growth factors is not enough long-term, and besides, is not without serious side effects. From this perspective, the greatest opportunities is the development of techniques using natural pool of pluripotent cells with high angiogenic potential

Hospital epidemiology of abdominal adhesions

The purpose of the research was to establish main causes of development and outcomes of adhesive obstruction. We analyzed the outcomes of treatment of 154 patients with acute intestinal obstruction. The following data were taken into consideration: age, gender, surgeries in past medical history, terms of adhesions manifestations, clinical symptoms, findings of additional examinations, methods of treatment, outcome. It has been established, that the most frequent causes of adhesions were appendectomy (23 %), stomach operations (21 %) and gynecological interventions (14 %). After surgeries performed in conditions of regional hospitals, adhesions developed after appendectomy, gynecological operations and interventions on the intestine. In patients operated in city hospitals intestinal obstruction more often occurred after extensive operations on the stomach, appendix, and after gynecological interventions. Urgent surgical interventions more often led to adhesive process in the abdominal cavity (63 %). Mortality made 5.8 %. Causes of death were intestinal fistulas and abdominal sepsis. Thus, acute adhesive intestinal obstruction as an extreme form of abdominal adhesions continues to be an urgent medical issue

Matrix metalloprotease 9 and remodeling after myocardial infarction

The objective of the work was to study the distribution of metalloprotease 9 (MMP9) in tissues after myocardial infarction. We studied anatomopathological material from 30 patients, the primary victims of acute transmural myocardial infarction. Average age - 63,7 years. There were 18 men and 12 women. 16 patients died in the period up to 3 days, 14 patients - in a period from 3 days to 1 month. Samples were fixed in formalin and immunohistochemical staining for MMP9 was conducted. We used rabbit IgG monoclonal antibody to MMP9 (Epitomics, Clone 1D: EP1254, Cat. N 2551-1, Lot YG 113001P) in the working dilution of 1: 100 - 1: 250. At fatal outcomes that happened within a few hours after the onset of a heart attack a large number of MMP9 was detected in neutrophils in the blood vessels at near infarction zone. In the period from 1 to 2 hours MMP9 was detected in the extracellular matrix in the area of infarction. In the period from 3 to 30 days MMP9 was detected only in fibroclasts at forming cardiosclerosis zone. Thus, the use of staining for MMP9 is convenient for determination of the time elapsed after myocardial infarction in post mortem examination

Expression of endothelin in experimental myocardial infarction in a changed concentration of FGF and VEGF

The objective of the work was to study the expression of endothelin in experimental myocardial infarction, and. to assess the impact on its altered concentrations of growth factors. The study was performed on a model of myocardial infarction. The experiment was conducted, on 150 female Wistar rats. Five groups of animals were studied: control group, the increased concentration FGF2 and. VEGF, the decreased concentration of FGF or VEGF. Excretion of animal experiments were conducted in the period from 1 to 30 days. The heart of the experimental animal was fixed in a solution of 10% neutral formalin. His-tochemical staining was used. Primary antibodies to endothelin ET-1/2/3 (H-38) rabbit polyclonal IgG (Santa Cruz, Cat. N Sc-98727, Lot # G2209). We determined that in control group there was a stain on endothelin in the area of myocardial infarction from the 1st to the 7th day with a peak on day 3. The use of growth factors increased the staining of the border and. infarct zones on the 1st to the 14th days with a maximum of day 3. The use of antibodies to growth factors reduced the stain of the in farcted area early after myocardial infarction (days 1 to 3), to the 7th day of the stain intensity increases sharply, reaching a maximum, and. to the 14th day again declined. Thus, we established the dynamics of expression of endothelin in the myocardium when the ischemic injury and. heart attack, as well as the significant stimulatory effect of growth factors FGF2 and. VEGF on the expression of endothelin in the area of ischemic damage at experimental myocardial infarction