[Effect of interleukin -27 on recovery from experimental autoimmune encephalomyelitis in C57BL/6 mice]

Multiple sclerosis and its murine model, experimental autoimmune encephalomyelitis [EAE], are chronic inflammatory demyelinating diseases of CNS. This study aimed to examine the effects of IL-27coding plasmid on disease status and certain immunological parameters in EAE-affected C57BL/6 mice. IL-27 gene was subcloned in P240 plasmid. The recombinant P240-mIL27 and P240 plasmids were injected two times, each time 200 micrograms, to test and control EAE mice, respectively. The clinical signs of the treated mice were evaluated daily and scored according to a standard method. One week after the last injection, all mice were sacrificed. The ELISA and MTT tests were performed to evaluate the production of IL-4, IFN-? and IL-17 from and proliferative response of splenocytes against specific antigen challenge, respectively. Furthermore, to demonstrate the immune cells infiltration, histopathological exam was performed on the brainstem of mice. The P240-mIL27 plasmid could significantly improve clinical course of EAE in the test group. Also in this group, the level of IL-4 was greater than that in the control group, while the levels of IFN-? and IL-17 were lower than those of the control group. In MTT test, the splenocytes of the test group showed a significantly less proliferative response than the control group. Finally, less infiltration of immune cells was seen in the brainstem of EAE mice treated with P240-mIL27 plasmid. IL-27 by shifting the immune responses from inflammatory Th1/Th17 towards anti-inflammatory Th2-type responses could be a suitable candidate for the treatment of inflammatory diseases such as MS

[Induction of allogeneic subcutaneous glioma tumor with GL 26 cell line in balb/c mice]

Glioma is the most common primary brain tumor. Despite many advances in treatment, all patients die within 6 to 18 months after diagnosis. In the cases of glioma, the immune system is suppressed in a local fashion. Therefore, unveiling the cellular and molecular mechanisms involved, with the aim of obtaining an appropriate new treatment is a priority. Designing an appropriate animal model is necessary before any clinical trials. In this study, we prepared fifteen 6-8 week-old female mice [Balb/C] from the Pasteur institute, Tehran, and also selected the mouse glioma cell line GL26 to induce a allogeneic subcutaneous tumor. After culturing the cell and anesthetization of the mice, we injected different cell doses into distinct groups of mice. Sterile PBS was injected into the control group. Animal behavior and clinical symptoms were regularly followed and recorded, and after tumor induction, it was surgically removed and evaluated in terms of macroscopic and microscopic characteristics. The tumor was induced more quickly with higher number of GL26 cells in mice. Atrophy and weakness was observed in the affected animals. In macroscopic examination, the tumor was relatively large, thick and full of blood. Moreover, in microscopic examination, cell proliferation, mitosis, abundant vessels, and tumor necrosis were observed. Regarding the limitations of a glioma syngeneic animal model, establishment of an allogeneic subcutaneous model, allows an easy evaluation of the size and volume of the tumor, without a requirement for sacrificing the animal. This model has the potential to provide opportunities for research on some immunological parameters, the testing of new therapeutic agents, and new discoveries in basic research, concerning glioma, for the first time in Iran

Time-dependent changes of immunologic responses after burn injury and immunomodulation by cimetidine and pyrimethamine in an animal model

Severe suppression of the immune system is the major cause of infections following burn injury. The aim of this study was to investigate the time-related alterations of immune responses following thermal injury in an animal model and also to modulate immune responses by use of the immunomdulators cimetidine and pyrimethamine. Male Balb/c mice were anesthetized and given a 10% total body surface area full-thickness burn. The timedependent changes of delayed type hypersensitivity [DTH] and antibody responses to sheep red blood cell [SRBC] were assessed at post-burn days [PBD]. The effects of different doses of cimetidine and pyrimethamine on DTH response were also quantitated at 10 PBD. Marked suppression of DTH response occurred during 30 days after burn trauma, with maximal suppression occurring between 10 to 14 days after burn injury. Simultaneously the antibody response to SRBC was significantly increased after thermal trauma. Cimetidine [at doses of 10 and 15 mg/kg] and pyrimethamine [at doses of 5 and 10 mg/kg] significantly augmented DTH response after thermal injury. These results showed that the severe time-dependent alterations occurred in DTH and antibody responses following burn injury. Cimetidine and pyrimethamine also restore burn-induced suppression of DTH response following thermal trauma

Kinetics of nitric oxide production and MTT reduction by HSV-1 infected macrophages

Macrophages have important role in defense against Herpes Simplex Virus type-1 [HSV-1]. The present study was performed to determine the viability and nitric oxide [NO] production by HSV-1 infected mouse peritoneal macrophages [HIM]. The viability of macrophages was evaluated using MTT reduction assay and the production of nitrite using Griess method. The ability of infected macrophages to reduce Tetrazolium [MTT] was diminished at virus to cell ratios of multiplicity of infection [MOI] of one, three and 10; but not at 0.01 and 0.1. Induction and inhibition of NO production by HIM were MOI dependent. The basal NO production by these cells was inhibited at MOI of three and ten. In contrast virus to cell ratios of 0.01 and 0.1 induced low but significant enhancement in NO production. The inability of HIM to reduce MTT at MOI of three was significant after 12-hrs and inhibition of NO production was initiated between 12-20 hours after infection. High doses of HSV-1 seem to decrease the normal activity of macrophages by inhibiting the production of nitric oxide

cytotoxicity pathway of natural killer cells in cord blood compared to peripheral blood

The cytotoxic activity of natural killer cells is usually tested by radioactive assay [51Cr release assay], which detects the release of cytoplasmic contents after plasma membrane disintegration of dying cells. In contrast to this indirect evaluation of cytotoxicity, the assessment of cell damage by flow cytometry aims to provide a more exact characterization of the death pathway via detection of the percentage of apoptosis and necrotic cells. Annexin V-FITC [Axv -FITC] can be used to label cells in the early apoptopic state, while propidium iodide [PI] indicates late apoptosis or necrosis. The NK cytotoxicity of cord blood [CB] and peripheral blood [PB] was determined after 4 hours of incubation in the absence of cytokines. After 4 hours in vitro incubation, co-staining with Annexin V-FITC [Axv-FITC] and propidium iodide [PI] permitted discrimination between viable, early apoptotic and necrotic cells. As we would expect, the cytotoxicity pathway in PB mononuclear cells [MNCs] consists of both apoptosis and necrosis pathways but in CB MNCs it almost consists of early apoptosis; and necrosis is negligible. With escalating E: T [effector: target] ratio changes in the percentage of apoptotic cells in PB samples were significantly higher than CB samples. The mechanism [s] of the low cytotoxicity of resting cord NK cells is not well understood. Complementary research in this field is recognized to elucidate the phenotypical and functional properties of CB cells and how they relate to maturational stages. CB studies are important for transplantation research and may provide insight to the suppressive mechanism by which the host-recipient could evade GVHD and rejection