A novel activity on thymocytes cells exerted by the rattlesnake (Crotalus durissus cumanensis) venom / Nueva actividad de los timocitos estimulada por el veneno de la serpiente de cascabel (Crotalus durissus cumanensis)

Abstract. Introduction: The thymus is active mainly during the neonatal and pre-adolescent periods. Objective: To test naïve thymocytes proliferation and monocytes stimulation. Materials and methods: We collected fresh thymus tissue from neonate mice after surgery. Suspension cells were coated onto Ficoll-Hypaque support. The obtained cells (thymocytes) were cultured measuring the proliferation of naïve T cells stimulated by Crotalus durissus cumanensis (Cdc) venom at sub-lethal doses (20 ng). Then, we supplemented the wells with AlamarBlue™ and incubated them for 5 h to test their proliferation. Mononuclear cells from mice peripheral blood were collected and layered onto the support of the Ficoll-Hypaque solution. We added the thymocytes actively dividing (25 x 105 cells) from cultures stimulated with Cdc venom at 20 ng/well to cultured monocytes freshly obtained from the Ficoll-Hypaque separation. Both cell populations were incubated for 36 h until monocytes matured to macrophages. Results: The naïve thymocytes rapidly proliferated after stimulation with the Cdc venom (NTCdc) and these successively induced the maturation and function of monocytes progenitor cells to mature macrophages, which ingested Chinese ink. Conclusions: The naïve thymocytes proliferated by stimulation with the Cdc venom and subsequently the NT/Cdc induced the rapid maturation and function of monocytes progenitor cells becoming mature macrophages with their phenotypic characteristics.

Resumen. Introducción. El timo es activo principalmente durante los períodos neonatal y preadolescente. Objetivo. Probar la proliferación de los timocitos tempranos y la estimulación de monocitos que producen. Materiales y métodos. Se recogió tejido de timo fresco después de la cirugía de ratones recién nacidos. La suspensión de células se colocó sobre un soporte de Ficoll-Hypaque. Las células obtenidas (timocitos) se cultivaron y se midió la proliferación de células T vírgenes estimuladas por el veneno de Crotalus durissus cumanensis (Cdc) en dosis subletales (20 ng). A continuación, se agregó AlamarBlue™ a los pocillos y se incubaron durante 5 horas para evaluar la proliferación. Se recogieron células mononucleares de sangre periférica de ratones y se colocaron sobre un soporte de solución de Ficoll-Hypaque. Los timocitos que se dividieron activamente (25 x 105 células) a partir de los cultivos estimulados con veneno de Cdc (20 ng/pocillo) y se agregaron a los cultivos de monocitos recién obtenidos de la separación en la solución de Ficoll-Hypaque. Ambas poblaciones celulares se incubaron durante 36 horas hasta que los monocitos maduraron a macrófagos. Resultados Los timocitos tempranos experimentaron una rápida proliferación estimulada por el veneno de Cdc (NTCdc) y, posteriormente, indujeron la maduración y la función de las células progenitoras de monocitos, los cuales maduraron a macrófagos, que se tiñeron con tinta china. Conclusiones. Los timocitos tempranos proliferaron con la estimulación del veneno de Cdc y, posteriormente, el NT/Cdc indujo la maduración rápida y la función de las células progenitoras de monocitos, transformándose en macrófagos con sus características fenotípicas.

CD24⁺ Cell Depletion Permits Effective Enrichment of Thymic iNKT Cells While Preserving Their Subset Composition

Invariant NKT (iNKT) cells are a small subset of thymus-generated T cells that produce cytokines to control both innate and adaptive immunity. Because of their very low frequency in the thymus, in-depth characterization of iNKT cells can be facilitated by their enrichment from total thymocytes. Magnetic-activated cell sorting (MACS) of glycolipid antigen-loaded CD1d-tetramer-binding cells is a commonly used method to enrich iNKT cells. Surprisingly, we found that this procedure also dramatically altered the subset composition of enriched iNKT cells. As such, NKT2 lineage cells that express large amounts of the transcription factor promyelocytic leukemia zinc finger were markedly over-represented, while NKT1 lineage cells expressing the transcription factor T-bet were significantly reduced. To overcome this limitation, here, we tested magnetic-activated depletion of CD24⁺ immature thymocytes as an alternative method to enrich iNKT cells. We found that the overall recovery in iNKT cell numbers did not differ between these 2 methods. However, enrichment by CD24⁺ cell depletion preserved the subset composition of iNKT cells in the thymus, and thus permitted accurate and reproducible analysis of thymic iNKT cells in further detail.

T Cell's Sense of Self: a Role of Self-Recognition in Shaping Functional Competence of Naïve T Cells

Post-thymic naïve T cells constitute a key cellular arm of adaptive immunity, with a well-known characteristic of the specificity and robustness of responses to cognate foreign antigens which is presented as a form of antigen-derived peptides bound to major histocompatibility complex (MHC) molecules by antigen-presenting cells (APCs). In a steady state, however, these cells are resting, quiescent in their activity, but must keep full ranges of functional integrity to mount rapid and robust immunity to cope with various infectious pathogens at any time and space. Such unique property of resting naïve T cells is not acquired in a default manner but rather requires an active mechanism. Although our understanding of exactly how this process occurs and what factors are involved remains incomplete, a particular role of self-recognition by T cells has grown greatly in recent years. In this brief review, we discuss recent data on how the interaction of T cells with self-peptide MHC ligands regulates their functional responsiveness and propose that variable strength of self-reactivity imposes distinctly different levels of functional competence and heterogeneity.

Protective and therapeutic impact of taurine on some biochemical, immunological and histological parameters in diabetic rats.

Background: Taurine is a non protein amino acid found in most animal tissues. It is a powerful antioxidant which shares in combating the harmful effect of the reactive oxygen species (ROS), associated to many chronic diseases as diabetes mellitus (DM). The disease is characterized by hyperglycemia and metabolic disorders in the body that leads to the release of ROS in the cells. Methods: The present work evaluates the biochemical and immunological role of taurine (500 mg/kg bwt) in ameliorating diabetes harm in rats when compared to the effect of the antidiabetic drug (amaryl). Six groups were established for the experiment. Group1: control rats without any supplementations. Group 2 : diabetic non treated rats. Group 3: rats received taurine for three weeks. Group 4: rats were supplemented with taurine for three weeks then injected streptozotocin (STZ) (prophylactic gp). Group 5: rats were injected with STZ then supplemented with taurine for four weeks (therapeutic gp). Group 6: rats were injected STZ then treated with amaryl drug for four weeks. Serum glucose and insulin levels in addition to liver function enzymes and lactate dehydrogenase enzyme were determined. ROS effect was monitored in liver tissue by detecting malondialdhyde resulting from lipid peroxidation and detecting glutathione reductase enzyme activity. With respect to the immunological responses, the thymocytes and splenocytes numbers were counted besides measuring serum IgG level. Histological and immunohistochemical studies were performed in pancreatic sections. Results: showed the ability of taurine in decreasing glucose level and increasing insulin with the same efficacy as amaryl drug besides affecting liver enzymes and improving the antioxidant system in cells. Taurine also restored the decrease in mean number of thymocytes and splenocytes caused by DM. Sera IgG levels from pre- and post-treatment with taurine showed non significant increase compared to the diabetic non treated group. Conclusion: post-treatment supplemention of taurine is recommended for T2DM.

Fibronectin modulates thymocyte-thymic epithelial cell interactions following Trypanosoma cruzi infection

Developing thymocytes interact with thymic epithelial cells (TECs) through cell-cell interactions, TEC-derived secretory moieties and extracellular matrix (ECM)-mediated interactions. These physiological interactions are crucial for normal thymocyte differentiation, but can be disrupted in pathological situations. Indeed, there is severe thymic atrophy in animals acutely infected with Trypanosoma cruzi due to CD4+CD8+ thymocyte depletion secondary to caspase-mediated apoptosis, together with changes in ECM deposition and thymocyte migration. We studied an in vitro model of TEC infection by T. cruzi and found that infected TEC cultures show a reduced number of cells, which was likely associated with decreased proliferative capacity, but not with increased cell death, as demonstrated by bromodeoxyuridine and annexin-V labelling. The infected TEC cultures exhibited increased expression of fibronectin (FN), laminin (LM) and type IV collagen. Importantly, treatment with FN increased the relative number of infected cells, whereas treatment with anti-FN or anti-LM antibodies resulted in lower infection rates. Consistent with these data, we observed increased thymocyte adhesion to infected TEC cultures. Overall, these results suggest that ECM molecules, particularly FN, facilitate infection of the thymic epithelium and that the consequent enhancement of ECM expression might be associated with changes in TEC-thymocyte interactions.

Expression of IL-17A during Thymus Regeneration in the Rat / 체질인류학회지

IL-17A is a pro-inflammatroy cytokine secreted by activated T cells. The IL-17 family consist of IL-17A, IL-17B, IL-17C, IL-17D, IL-17E and IL-17F. IL-17A and IL-17F are produced primarily in activated T cells. In contrast, IL-17B, IL-17C, IL-17D and IL-17E are expressed in a wide assortment of tissues. Their functions partially overlap those of IL-17A, although they have not been as thoroughly investigated. The receptor for IL-17A (IL-17R) is widely expressed in a variety of tissues. IL-17A and IL-17E mRNAs were expressed in only EL4 cells. IL-17C mRNA expression was observed in the thymic subcapsular/cortex epithelial cells (SNEC), cortex or cortical reticular cells (CREC), medullary epithelial cells (MEC), medullary interdigitating-like cells (MDC), thymocytes and EL4 cells. However, IL-17C mRNA was not expressed in RAW 264.7 cells. Immunohistochemical study also demonstrated not only the presence of IL-17A mainly in the thymic epithelial cells, but also the upregulated expression of IL-17A in the thymic epithelial cells of the regenerating thymus. Thus, the results of the present study suggest that IL-17A expressed in the thymocytes and thymic epithelial cells could play an important role in the development of new T cells to replace T cells damaged by cyclophosphamide treatment during thymus regeneration.

Protective effect of polypeptides from Chlamys farreri on murine thymocytes damaged by 60Co γ-rays / 中华放射医学与防护杂志

Objective To study the effect and mechanism of polypeptide from Chlamys farreri(PCF) on the thymocytes damaged by 60Co γ-rays. Methods The ceils were randomly divided into six groups: control group, 60Co groups (2 Gy, model),60Co + 0.5% PCF, 60Co+ 0.25% PCF, 60Co + 0.125% PCF, 60Co +0.1% VitC. The concentration of GSH-Px, ROS, A-SAC, T-AOC and the cells' viability were determined. The mitochondria membrane potential were tested. The expressions of P53, Bax and Bel-2 proteins were examined.Results The activities f GSH-Px, A-SAC, T-AOC in cells were enhanced, and the amounts of ROS were decreased by PCF. The expression of Bcl-2 gene was up-regulated, and down-regulated for the expression of P53 and Bax. All observed indexes of the PCF groups were significantly different compared with model group (P <0.05). Conclusions PCF has the protective effects on damages of thymocytes caused by 60Co irradiation. The mechanisms might be related to inhibiting of lymphocyte apoptosis and enhancing the activities of lymphocytes.

The mitochondrial and structural protein changes in dexamethasone-induced mouse thymocyte apoptosis / 中国病理生理杂志

AIM: To study mitochondrial mass and structural protein changes in dexamethasone (DEX)-mediated mouse thymocyte apoptosis process. METHODS: DEX-induced mouse thymocyte apoptosis model was established. Annexin V-FITC/PI double staining was used to identify apoptotic and necrotic cells by flowcytometry, JC-1 staining was adopted to test mitochondrial membrane potential (△?_m), and cellular structural protein changes were studied with CFDA-SE staining. RESULTS: By 1?10~(-6) mol/L DEX stimulation, the apoptotic rate was 51.25%?5.51% and had significantly difference from control group (12.03%?2.00%); the necrotic rate in DEX group was 30.25%?3.67% and also had significantly difference from control group (10.11%?1.11%, P

Expression of proteins in mouse thymocytes induced by low dose X-rays and their biological activity / 中国病理生理杂志

AIM: To observe the expression of proteins induced by whole-body irradiation in mice with low dose X-rays and their biological activity. METHODS: To analyse the expression of protein induced by whole-body irradiation with 75mGy X-rays and sham-irradiation, the method of gel filtration with Sephadex G-100 and HPLC were used. The biological activity of protein expressed in thymocytes was observed by mouse splenocyte proliferation and chromosome aberration of human peripheral blood lymphocytes. RESULTS: HPLC analysis showed that there was a marked decrease in expression of 24.5 kD protein molecule in the fractions of thymocyte extract in comparison with the corresponding fractions from the sham-irradiated control mice. These protein fractions from the thymocytes of the irradiated mice at the concentration of 6.25 mg/L showed both inhibitory effect on normal T cell proliferation and protective effect on the chromosome damage induced by high dose radiation. CONCLUSION: Down regulation of 24.5 kD protein molecule expression may have implications in the mechanism of immunoenhancement and cytogenetic adaptive response induced by low dose radiation.

The effects of PCF on the activity and recoveration of thymocytes irradiated by ~(60)Co / 中国海洋药物

In this paper the protective effect and facillitating reconverting effect of polypeptide from chlamys farreri (PCF) on thymocytes irradiated by 60 Co ? ray were studied using the MTT chromatometry .The results showed that:(1)the damaged extents of thymocytes were increased with the radiative intensity of 60 Co ? ray elevated at the range of 3GY to 9GY compared with that in no-exposure to 60 Co ? ray group.(2)PCF could reduce 60 Co ? ray damage on thymocytes with dose dependence .In the 0.25%~4% concentration range the higher concentration of PCF the stronger protective effects of it ,but the protective effect of PCF at the concentration below 0.5% disappeared while the radiative intensity of 60 Co ? ray was at 9GY.(3)PCF could facilitate the renovation ability of thymocytes after exposure 60 Co ? ray for two hours .And only 2% concentration of PCF showed the facilitating repair process of thymocytes after radiated by 60 Co ? ray for 7 hours .(4) PCF could decrease the activity and facilitate the death of the dying thymocytes after exposure 60 Co ? ray for 19 hours. The results suggested that the protective effect and facilitating repair effect of PCF on the thymocytes damaged by 60 Co ? ray may be mediated by the antisuperoxidation of PCF.

Diverse Expression of NK Cell Receptor between Fetal Thymocytes and Fetal Liver Lymphocytes from the Same Individuals / 대한면역학회지

Fetal thymus may be the organ for NK cell maturation, but the in vivo evidences are few, Here, by analyzing NK cell receptor, we present that NK cells develop in fetal thymus and fetal liver and that NK cell receptor appears earlier than the expression CD16 or CD56. Moreover, the finding that the repertoire of NK cell receptor is different between fetal thymus and fetal liver lymphocytes suggests that the environmental factors may influence the NK cell receptor repertoire during NK cell maturation.

Expression of p58 in Fetal Thymocytes and Fetal Liver Lymphocytes / 대한면역학회지

Bipotent progenitors for T and natural killer (NK) lymphocytes are thought to exist among early precursor thymocytes or liver lymphocytes. The identification of such a progenitor population or mature NK cells in such organs remains undefined. Here we report the identification of a novel receptor of NK cells, p58 (HLA class I-specific inhibitory receptors), in fetal thymocytes and fetal liver lymphocytes. Our finding suggests the NK cells mature in the developmental stage during feta1 ontogeny. Flow cytometric analysis revealed p58 positive cells in thymocytes or in fetal liver lymphocytes and reverse transcription PCR also showed amplification of p58 RNA. The result of single stranded conformational polymorphism (SSCP) showed it discriminates one or two base pair differences of the p58 gene. Although the question still remains as to whether the expression of p58 is due to the NK cells or natural T cells, it is clear the p58 is expressed in fetal thymocytes or liver lymphocytes. And SSCP analysis using appropriate sets of primers used in this study, is helpful to study the diversity of p58.

Protective effect of polypeptide from Chlamys farreri on UVB-irradiated murine thymocytes in vitro / 中国海洋药物

Objective To study the radioprotective effect of polypeptide from Chlamys farreri (PCF) on ultraviolet B(UVB)-irradiated murine thmocytes in vitro. Methods The murine thymocytes were exposed to UVB radiation. MTT method was used to detect the cell viability. The activities of intracellular glutathione peroxidase (GSH-Px), superoxided dismutase(SOD) and catalase (CAT) were measured. Intracellular reactive oxygen species (ROS) and the effect of PCF on UVB-induced apoptosis were investigated by flow cytometry. Results PCF could greatly enhance the viability of murine thymocyte and markedly promote the activities of GSH-Px, SOD and CAT, while the amount of ROS was decreased. PCF could inhibit UVB-induced thymocyte apoptosis. Conclusion PCF has significant radioprotective effects on UVB-irradiated murine thymocytes in vitro.

Study on the protective effect of polypeptide from Chlamys farreri(PCF) against murine thymocytes apoptosis induced by UVB radiation / 中国海洋药物

Objective To study the protective effect of PCF against murine thymocytes apoptosis induced by UVB irradiation.Methods The irradiation damage model of thymocytes caused by UVB was established.Cellular ultrastructuralalterations were observed with TEM.The expressions of P53,Bax and Bcl-2 proteins were examined by immunocytochemical technique. The p38 mRNA gene expression was also determined with the technique of in situ hybridization.Results Apoptotic thymocytes were observed after UVB irradiation.The ultrastructural damages of the cells induced by UVB irradiation were reduced by the treatment of PCF.PCF also raised the expression of Bcl-2 gene and decreased the expression of P53,Bax.The mRNA expressions of p38 were decreased in PCF groups.Conclusion PCF showed significant protective effects on UVB-irradiated murine thymocytes in vitro.

Changes of mitochondrial membrane potential in mouse thymocytes stimulated by dexamethasone / 中国病理生理杂志

0.05) compared with control group at 1 h and 3 h; while ~FL 1 in DEX group at 5 h (660.91?72.95) was significant lower (P

Effects of thymosin ?_1 on the development and maturation of thymocytes / 中国病理生理杂志

AIM: To investigate the effect of Thymosin ? 1 on the development and matutation of thymocytes. METHODS: The proportion of CD4+CD8+ thymocytes and the expression of smoothened (Smo) of the hedgehog (Hh)-signaling in CD4-CD8-thymocytes were examined to observe the effect of thymosin ? 1 on thymocyte development and matutation. RESULTS: Flowcytometric analysis showed that thymosin ? 1 showed activity at a low dose of 30 ?g/kg, and 30 ?g/kg thymosin ? 1 accelerated the replenishment and maturation of thymocytes according to the expression of Smo of the Hh-signaling in CD4-CD8-thymocytes, the potent negative regulator of proliferative responses. CONCLUSION: Thymosin ? 1 can accelerate thymocyte development from CD4-CD8- to CD4+CD8+.

Effects of Shaoqiduogan on cellular immunity in mice with delayed-type hypersensitivity / 中国药理学通报

Aim To investigate the effects of Shaoqiduogan(SQDG)on cellular immunity in mice with delayed-type hypersensitivity(DTH). Methods DTH model induced by 2, 4-dinitrochlorobenzene(DNCB)and cyclophosphamide(Cy)suppressed and potentiated DTH model were used. SQDG(30, 60 and 120 mg ? kg-1)were given intragastrically(ig)once daily for 7 consecutive days. Concanavalin A(Con A)-induced T cells proliferation was measured by MTT assay. The activity of IL-2 in culture supernatants of Con A-induced T cells proliferation was measured by testing its ability to support the proliferation of Con A stimulated thymocytes which was detected by MTT assay. Results In mice with DTH, SQDG(120 mg?kg-1)reduced ear swelling, thymus index, Con A-induced T cells proliferation and the activity of IL-2 in culture supernatants of Con A-induced T cells proliferation. The suppression and potentiation of DTH in mice were induced by given an intraperitoneal(ip)injection of Cy at the dose of 150mg?kg-1 on primary sensitization day and 250 mg?kg-1 three days before primary sensitization. SQDG(60 and 120 mg?kg-1)upregulated ear swelling, Con A-induced T cells proliferation and the activity of IL-2 in culture supernatants of T cells from mice with Cy-suppressed DTH; and downregulated levels of the aforementioned indexes in mice with Cy-potentiated DTH. Conclusion SQDG possesses a dual modulatory effect on cellular immunity in mice.

The feature of TCR D?X gene rearrangements in peripheral blood T cells and thymocytes from healthy individual / 中国免疫学杂志

Objective: To investigate the distribution feature of TCR D~X gene rearrangement in peripheral blood T cells and thymocytes from healthy individuals. Methods: The rearrangement segments of TCR D?X gene with J?X, D?3 or Ja were amplified in genomic DNA from peripheral blood mononuclear cells(PBMCs) of 10 cases, sorted CD3 + T cells from peripheral blood of 4 cases and thymocytes from 7 cases, by using semi-nested PCR. Different amounts of DNA from all samples were amplified to estimate the frequency of D?X gene rearrangements. Results: The rearrangements of TCR D?X with J?X,D?3 or PJa respectively could be found in the most samples of peripheral blood or thymocytes.The frequency of D?X rearrangements were different in peripheral blood cells and thymocytes by analysis of PCR with different amounts DNA. Conclusion: The TCR D?X, Ja was the most frequent rearrangement in mature and immature T cells, whereas TCR D?X,D?3 were more frequently rearranged in immature T cells.

Effect of ERK inhibition on the mitochondrial potential change in dexamethasone-induced thymocyte apoptosis / 中国病理生理杂志

AIM: To study the effect of ERK inhibition on the mitochondrial potential change in dexamethasone (DEX)-induced thymocyte apoptosis. METHODS: ERK activity was inhibited by PD098059 (PD), and 4 experimental groups were set: control, PD only, DEX and PD+DEX. Annexin V-FITC/PI double staining flowcytometry was used to detect apoptotic cells at time points of 3 h, 5 h and 7 h. JC-1 staining flowcytometry was adopted to examine mitochondrial membrane potential (△?m) at time points of 3 h, 7 h and 11 h. RESULTS: By stimulation with 1 ?mol/L DEX, the apoptotic rates of mouse thymocytes at 3 h, 5 h and 7 h were (19.63?0.35)%, (41.84?1.67)% and (67.00?2.43)%, respectively, and had significantly difference from control group (4.98?0.39)%, (6.08?0.33)% and (9.31?0.34)% (P0.05). At 3 h, 7 h and 11 h, the rates of low △?m cells were (21.23?1.43)%, (55.34?1.78)% and (70.88?2.87)%, significantly higher than that in control group (P0.05). CONCLUSION: DEX induces mouse thymocyte apoptosis at least partly through ERK pathway, and ERK inhibition has an important biological significance during this process.

The c-Myc expression and apoptosis of mouse thymocytes treated with dexamethasone / 中国病理生理杂志

AIM: To study c-Myc expression and its relationship with caspase-3 in a dexamethasone (DEX)- induced mouse thymocyte apoptosis model, and discuss the role of c-Myc in cell apoptosis. METHODS: Mouse thymocyte apoptosis was induced by 1 ?mol/L DEX, the apoptotic and necrosis cells were measured by Annexin V-FITC/PI double staining flowcytometry at 30 min, 3 h, 6 h and 9 h . Electron microscopy observation was carried out at 6 h, and c-Myc and caspase-3 contents were tested by Western blot at 0, 30, 60, 180 min. RESULTS: By 1?mol/L DEX treatment, the apoptosis rates of thymocytes at 30 min, 3 h, 6 h, 9 h were (5.70?0.46)%, (35.79?1.13)%, (50.61?2.15)% and (35.52?1.66)%,respectively; in control group, they were (5.97?0.25)%, (10.20?0.71)%, (12.10?0.66)% and (15.45?0.51)% (P