Interleukin-21 expanded NKDC in vitro reduces the B16F10 tumor growth in vivo

Innate immunity to tumors is mediated mainly by natural killer cells (NKs) and dendritic cells (DCs). The function of these cells is coordinated by cytokines produced during the inflammatory process. NK cells are highly active against tumors, being an important source of IFN-γ. Natural killer dendritic cells (NKDCs) were recently identified as a group of hybrid cells; some studies claim that they have lytic activity, produce IFN-γ and can also stimulate antigen-specific T cells. Interleukin 21 (IL-21) regulates the proliferation capacity and cytotoxicity of NK and T cells. The main objective of this study was to investigate if IL-21 influences the frequency of NKDCs in vitro as well as IFN-γ production and also to verify if these cells could enhance the antitumor activity against B16F10 tumor model in vivo. Splenocytes from C57BL/6 mice were isolated and the DC were enriched by immunomagnetic beads and cultured for four days with recombinant IL-21 (10, 20, 40 or 100 ng/ml). NKDC population was characterized as CD11c^low/medB220⁺NK1.1⁺. Expanded cells were used to treat B16F10 tumor bearing mice and tumor growth was compared between the doses of IL-21 10 ng/ml and 20 ng/ml. The results indicate that IL-21 increases the expansion of splenic NKDCs in vitro in doses of 10 ng/ml and 20 ng/ml and these cells produce IFN-γ. In vivo, cells expanded with IL-21 and injected directly into the growing tumor efficiently reduced the tumor size. Together, these results showed for the first time that IL-21 influences the biology and the effector activity of NKDCs.     

Similar hypotensive responses to resistance exercise with and without blood flow restriction

Low intensity resistance exercise (RE) with blood flow restriction (BFR) has gained attention in the literature due to the beneficial effects on functional and morphological variables, similar to those observed during traditional RE without BFR, while the effects of BFR on post-exercise hypotension remain unclear. The aim of the present study was to compare the blood pressure (BP) response of trained normotensive individuals to RE with and without BFR. In this cross-over randomized trial, eight male subjects (23.8 ± 4 years, 74 ± 3 kg, 174 ± 4 cm) completed two exercise protocols: traditional RE (3 x 10 repetitions at 70% one-repetition maximum [1-RM]) and low intensity RE (3 x 15 repetitions at 20% 1-RM) with BFR. Blood pressure measurements were performed after 15 min of seated rest (0), immediately after and 10 min, 20 min, 30 min, 40 min, 50 min and 60 min after the experimental sessions. Similar hypotensive effects for systolic BP (SBP) were observed for both protocols (P < 0.05) after exercise, with no differences between groups (P > 0.05) and no statistically significant difference for diastolic BP (P > 0.05). These results suggest that in normotensive trained individuals, both traditional RE and RE with BFR induce hypotension for SBP, which is important to prevent cardiovascular disturbances.     

Myiasis by screw worm Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae) in a wild maned wolf Chrysocyon brachyurus (Mammalia: Canidae), in Brasília, Brazil

In April 2009, a wild maned wolf, Chrysocyon brachyurus, was captured in an area of cerrado in Brasília, DF, Brazil, with screw worm maggots in external wounds. Fifty larvae were bred in the laboratory and eight adults of Cochliomyia hominivorax (Coquerel) emerged 10 days after pupation. This is the first report of a myiasis by C. hominivorax in a free-living maned wolf in Brazil.     

HspBP1 levels are elevated in breast tumor tissue and inversely related to tumor aggressiveness

HspBP1 is a co-chaperone that binds to and regulates the chaperone Hsp70 (Hsp70 is used to refer to HSPA1A and HSPA1B). Hsp70 is known to be elevated in breast tumor tissue, therefore the purpose of these studies was to quantify the expression of HspBP1 in primary breast tumors and in serum of these patients with a follow-up analysis after 6 to 7 years. Levels of HspBP1, Hsp70, and anti-HspBP1 antibodies in sera of breast cancer patients and healthy individuals were measured by enzyme-linked immunosorbent assay. Expression of HspBP1 was quantified from biopsies of tumor and normal breast tissue by Western blot analysis. The data obtained were analyzed for association with tumor aggressiveness markers and with patient outcome. The levels of HspBP1 and Hsp70 were significantly higher in sera of patients compared to sera of healthy individuals. HspBP1 antibodies did not differ significantly between groups. HspBP1 levels were significantly higher in tumor (14.46 ng/μg protein, n = 51) compared to normal adjacent tissue (3.17 ng/μg protein, n = 41, p < 0.001). Expression of HspBP1 was significantly lower in patients with lymph node metastasis and positive for estrogen receptors. HspBP1 levels were also significantly lower in patients with a higher incidence of metastasis and death following a 6 to 7-year follow-up. The HspBP1/Hsp70 molar ratio was not associated with the prognostic markers analyzed. Our results indicate that low HspBP1 expression could be a candidate tumor aggressiveness marker. This work was supported by FAPERGS, CNPq, and the National Institute of General Medical Sciences grant number GM072628-02 (V.G.) The expression of HspBP1 (an Hsp70 co-chaperone) was analyzed in tumor samples and sera from breast cancer patients. HspBP1 is over expressed in these tumors and a seven year follow-up analysis found an association with a poor prognosis. Chaperones have been shown to play important roles in tumor biology and immunology; therefore, we believe the data in this study will serve as a basis for the formulation of a new hypothesis on chaperone-co-chaperone interactions and their role in tumor growth.     

CD4<Superscript>+</Superscript> T cell response against a non-tumor antigen is unaffected in melanoma-bearing mice

The tumor microenvironment is complex and creates an immunosuppressive network to tolerize tumor-specific immune responses; however, little information is available regarding the response against non-tumor antigens in tumor-bearing individuals. The goal of the present study was to evaluate if tumor burden could influence a CD4⁺ T cell response against a soluble protein, not expressed by the tumor, in the absence of in vitro stimulation. Using an experimental system in which we can compare CD4⁺ T cell responses to the Ea antigen when it is either expressed by B16F10 melanoma cells (B16EaRFP cells) or is an exogenous, non-tumor antigen (soluble EaRFP protein), in immunizations of B16F10 tumor-bearing mice, we observed that the tumor can modulate the CD4⁺ T cell-specific response to the antigen when it is expressed by the tumor cells. TEa cells proliferated poorly and produced less IFN-γ in mice bearing B16F10 melanoma expressing Ea peptide, and tumor growth was impervious to this response. However, in mice bearing 7 days B16F10 tumors, not expressing the Ea antigen, priming of TEa cells was similar to that observed in tumor-free mice, based on the total number of cells recovered and proliferation assessed by CFSE dilution after EaRFP immunization. We also investigated if tumor burden could influence recall responses of already differentiated effector cells. We immunized mice with EaRFP antigen and after a few days injected B16F10 cells. After 10 days of tumor growth, we challenged the mice with the non-tumor antigen. We found that the number of TEa cells producing IFN-γ in tumor-bearing mice was not different compared to tumor-free mice. No differences in antigen presentation, assessed by YAe antibody staining, were verified in the draining lymph node of these two groups. Collectively, our data indicate that tumor burden does not affect immune responses to non-tumor antigens. These results have important implications in the design of anti-cancer therapy.     

Self-tolerance checkpoints in CD4 T cells specific for a peptide derived from the B cell antigen receptor

Linked recognition of Ag by B and T lymphocytes is ensured in part by a state of tolerance acquired by CD4 T cells to germline-encoded sequences within the B cell Ag receptor (BCR). We sought to determine how such tolerance is attained when a peptide from the BCR variable (V) region is expressed by small numbers of B cells as it is in the physiological state. Mixed bone marrow (BM) chimeras were generated using donor BM from mice with B cells that expressed a transgene (Tg)-encoded κ L chain and BM from TCR Tg mice in which the CD4 T cells (CA30) were specific for a Vκ peptide encoded by the κTg. In chimeras where few B cells express the κTg, many CA30 cells were deleted in the thymus. However, a substantial fraction survived to the CD4 single-positive stage. Among single-positive CA30 thymocytes, few reached maturity and migrated to the periphery. Maturation was strongly associated with, and likely promoted by, expression of an endogenous TCR α-chain. CD4(+) CA30 cells that reached peripheral lymphoid tissues were Ag-experienced and anergic, and some developed into regulatory cells. These findings reveal several checkpoints and mechanisms that enforce a state of self-tolerance in developing T cells specific for BCR V region sequences, thus ensuring that T cell help to B cells occurs through linked recognition of foreign Ag.