IKKα is required for the homeostasis of regulatory T cells and for the expansion of both regulatory and effector CD4 T cells.

It was reported that TNF receptor type II signaling, which has the capacity to stimulate CD4+ forkhead box P3+ (Foxp3+) regulatory T cells (Tregs), activated the noncanonical NF-κB pathway in an IKKα-dependent manner. Therefore, we studied the role of IKKα in the homeostasis of Treg population. To this end, we generated a mouse strain with conditional knockout of IKKα in CD4 cells (Ikkα(f/f):CD4.Cre) that showed a >60% reduction in the number of Tregs in the thymus and peripheral lymphoid tissues, whereas the number of Foxp3- effector T cells (Teffs) remained at a normal level. The function of Tregs deficient in IKKα was examined using Rag1(-/-) mice cotransferred with naive CD4 cells (nCD4s). Although wild-type (WT) Tregs inhibited colitis induced by transfer of WT nCD4s, IKKα-deficient Tregs failed to do so, which was associated with their inability to reconstitute Rag1(-/-) mice. Furthermore, nCD4s deficient in IKKα also failed to reconstitute Rag1(-/-) mice and were defective in proliferative responses in vitro and in vivo. Thus, our study reveals a novel role of IKKα in the maintenance of a normal Treg population and in the control of expansion of CD4 T cells. These properties of IKKα may be exploited as therapeutic strategies in the treatment of major human diseases.

TNFR2 is critical for the stabilization of the CD4+Foxp3+ regulatory T. cell phenotype in the inflammatory environment.

Several lines of evidence indicate the instability of CD4(+)Foxp3(+) regulatory T cells (Tregs). We have therefore investigated means of promoting the stability of Tregs. In this study, we found that the proportion of Tregs in mouse strains deficient in TNFR2 or its ligands was reduced in the thymus and peripheral lymphoid tissues, suggesting a potential role of TNFR2 in promoting the sustained expression of Foxp3. We observed that upon in vitro activation with plate-bound anti-CD3 Ab and soluble anti-CD28 Ab, Foxp3 expression by highly purified mouse Tregs was markedly downregulated. Importantly, TNF partially abrogated this effect of TCR stimulation and stabilized Foxp3 expression. This effect of TNF was blocked by anti-TNFR2 Ab, but not by anti-TNFR1 Ab. Furthermore, TNF was not able to maintain Foxp3 expression by TNFR2-deficient Tregs. In a mouse colitis model induced by transfer of naive CD4 cells into Rag1(-/-) mice, the disease could be inhibited by cotransfer of wild-type Tregs, but not by cotransfer of TNFR2-deficient Tregs. Furthermore, in the lamina propria of the colitis model, most wild-type Tregs maintained Foxp3 expression. In contrast, an increased number of TNFR2-deficient Tregs lost Foxp3 expression. Thus, our data clearly show that TNFR2 is critical for the phenotypic and functional stability of Tregs in the inflammatory environment. This effect of TNF should be taken into account when designing future therapy of autoimmunity and graft-versus-host disease by using TNF inhibitors.

ß-Defensin 2 and 3 promote the uptake of self or CpG DNA, enhance IFN-α production by human plasmacytoid dendritic cells, and promote inflammation.

Alarmins are a group of structurally diverse host defense antimicrobial peptides that are important immune activators. In this article, we present a novel role for two potent alarmins, human ß-defensin 2 and 3 (HBD2 and 3), in promoting IFN-α production by human plasmacytoid dendritic cells. We demonstrate that HBD2 and 3 activate pDCs by enhancing the intracellular uptake of CpG and self DNA and promote DNA-induced IFN-α production in a TLR9-dependent manner. Both CpG and host DNA form aggregates that resemble DNA nets when combined with HBD2 and 3. Isothermal titration calorimetry studies to elucidate the nature of HBD3/CpG complexes demonstrate involvement of enthalpy-driven interactions, in addition to hydrophobic interactions, with the formation of complexes at a molar ratio of 2:1 defensin/CpG. The i.v. administration of HBD3/CpG complexes induced proinflammatory cytokines like IL-12, IFN-γ, IL-6, IFN-α, and IL-10 in serum, associated with an increased recruitment of APCs in the spleen. Subcutaneous injections of these complexes showed enhanced infiltration of inflammatory cells at the injection site, indicating a potential pathophysiological role for alarmin/DNA complexes in contributing to inflammation. Intraperitoneal immunization of HBD3/CpG complexes with OVA enhanced both cellular and humoral responses to OVA, compared with OVA/HBD3 or OVA/CPG alone, indicative of a much more potent adjuvant effect of the HBD3/CpG complexes. Thus, the ability of defensins to enhance cellular uptake of nucleic acids can lead to improved vaccine formulations by promoting their uptake by various cells, resulting in an enhanced immune response.

Inducible NOS-induced chloride intracellular channel 4 (CLIC4) nuclear translocation regulates macrophage deactivation.

Nuclear translocation of cytosolic CLIC4 is an essential feature of its proapoptotic and prodifferentiation functions. Here we demonstrate that CLIC4 is induced concurrently with inducible nitric oxide synthase (iNOS) and S-nitrosylated in proinflammatory peritoneal macrophages. Chemical inhibition or genetic ablation of iNOS inhibits S-nitrosylation and nuclear translocation of CLIC4. In macrophages, iNOS-induced nuclear CLIC4 coincides with the pro- to anti-inflammatory transition of the cells because IL-1ß and CXCL1 mRNA remain elevated in CLIC4 and iNOS knockout macrophages at late time points, whereas TNFα mRNA is elevated only in the iNOS knockout macrophages. Active IL-1ß remains elevated in CLIC4 knockout macrophages and in macrophages in which CLIC4 nuclear translocation is prevented by the NOS inhibitor l-NAME. Moreover, overexpression of nuclear-targeted CLIC4 down-regulates IL-1ß in stimulated macrophages. In mice, genetically null for CLIC4, the number of phagocytosing macrophages stimulated by LPS is reduced. Thus, iNOS-induced nuclear CLIC4 is an essential part of the macrophage deactivation program.

Alarmin-induced cell migration.

Alarmins are endogenous, constitutively available, damage-associated molecular patterns that upon release can mobilize and activate various leukocytes for the induction of innate and adaptive immune responses. For our immune system to function appropriately, it relies on navigating various leukocytes to distinct places at the right time. The direction of cell migration is determined by chemotactic factors that include classical chemoattractants, chemokines, certain growth factors, and alarmins. This viewpoint provides an overview of alarmin-induced cell migration. Alarmins are capable of inducing the migration of diverse types of leukocytes and nonleukocytes either directly by triggering specific receptors or indirectly by inducing production of chemokines through the activation of various leukocytes via pattern recognition receptors. The receptors used by alarmins to directly induce cell migration can either be Gαi protein-coupled receptors or receptors such as the receptor for advanced glycation end products; however, the intracellular signaling events responsible for the direct chemotactic activities of alarmins are, to date, only partially elucidated. Given that alarmins act in concert with chemokines to regulate the recruitment and trafficking of leukocytes, these damage-associated molecular patterns are potentially involved in diverse biological processes as discussed in this viewpoint.

In vitro generated Th17 cells support the expansion and phenotypic stability of CD4(+)Foxp3(+) regulatory T cells in vivo.

CD4(+) T cells stimulate immune responses through distinct patterns of cytokine produced by Th1, Th2 or Th17 cells, or inhibit immune responses through Foxp3-expressing regulatory T cells (Tregs). Paradoxically, effector T cells were recently shown to activate Tregs, however, it remains unclear which Th subset is responsible for this effect. In this study, we found that Th17 cells expressed the highest levels of TNF among in vitro generated Th subsets, and most potently promoted expansion and stabilized Foxp3 expression by Tregs when co-transferred into Rag1(-/-) mice. Both TNF and IL-2 produced by Th17 cells contributed to this effect. The stimulatory effect of Th17 cells on Tregs was largely abolished when co-transferred with TNFR2-deficient Tregs. Furthermore, Tregs deficient in TNFR2 also supported a much lower production of IL-17A and TNF expression by co-transferred Th17 cells. Thus, our data indicate that the TNF-TNFR2 pathway plays a crucial role in the reciprocal stimulatory effect of Th17 cells and Tregs. This bidirectional interaction should be taken into account when designing therapy targeting Th17 cells, Tregs, TNF and TNFR2.

Structural plasticity of a transmembrane peptide allows self-assembly into biologically active nanoparticles.

Significant efforts have been devoted to the development of nanoparticular delivering systems targeting tumors. However, clinical application of nanoparticles is hampered by insufficient size homogeneity, difficulties in reproducible synthesis and manufacturing, frequent high uptake in the liver, systemic toxicity of the carriers (particularly for inorganic nanoparticles), and insufficient selectivity for tumor cells. We have found that properly modified synthetic analogs of transmembrane domains of membrane proteins can self-assemble into remarkably uniform spherical nanoparticles with innate biological activity. Self-assembly is driven by a structural transition of the peptide that adopts predominantly a beta-hairpin conformation in aqueous solutions, but folds into an alpha-helix upon spontaneous fusion of the nanoparticles with cell membrane. A 24-amino acid peptide corresponding to the second transmembrane helix of the CXCR4 forms self-assembled particles that inhibit CXCR4 function in vitro and hamper CXCR4-dependent tumor metastasis in vivo. Furthermore, such nanoparticles can encapsulate hydrophobic drugs, thus providing a delivery system with the potential for dual biological activity.

Painful pathways induced by TLR stimulation of dorsal root ganglion neurons.

We hypothesize that innate immune signals from infectious organisms and/or injured tissues may activate peripheral neuronal pain signals. In this study, we demonstrated that TLRs 3, 7, and 9 are expressed by human dorsal root ganglion neurons (DRGNs) and in cultures of primary mouse DRGNs. Stimulation of murine DRGNs with TLR ligands induced expression and production of proinflammatory chemokines and cytokines CCL5 (RANTES), CXCL10 (IP-10), IL-1α, IL-1ß, and PGE(2), which have previously been shown to augment pain. Further, TLR ligands upregulated the expression of a nociceptive receptor, transient receptor potential vanilloid type 1 (TRPV1), and enhanced calcium flux by TRPV1-expressing DRGNs. Using a tumor-induced temperature sensitivity model, we showed that in vivo administration of a TLR9 antagonist, known as a suppressive oligodeoxynucleotide, blocked tumor-induced temperature sensitivity. Taken together, these data indicate that stimulation of peripheral neurons by TLR ligands can induce nerve pain.

Expression of costimulatory TNFR2 induces resistance of CD4+FoxP3- conventional T cells to suppression by CD4+FoxP3+ regulatory T cells.

Our previous study showed that TNFR2 is preferentially expressed by CD4(+)FoxP3(+) regulatory T cells (Tregs), and expression of this receptor identified maximally suppressive Tregs. TNFR2 is also expressed by a small fraction of CD4(+)FoxP3(-) conventional T cells (Tconvs) in normal mice, and its expression is upregulated by T cell activation. This raises questions about the role of TNFR2 signaling in the function of Tconv cells. In this study, by using FoxP3/gfp knock-in mice, we showed that TNFR2 signaling did not induce FoxP3(-) CD4 cells to become suppressive. Ki-67, a marker of proliferation, was concomitantly expressed with TNFR2 by CD4 cells, independent of forkhead box P3 expression, in normal mice and Lewis lung carcinoma-bearing mice. TNFR2 is associated with greater suppressive functions when expressed by Tregs and is associated with greater resistance to suppression when expressed by Tconv cells. In mice bearing 4T1 breast tumor or Lewis lung carcinoma, intratumoral Tconv cells expressing elevated levels of TNFR2 acquired the capacity to resist suppression by lymph node-derived Tregs. However, they remained susceptible to inhibition by more suppressive tumor-infiltrating Tregs, which expressed higher levels of TNFR2. Our data indicate that TNFR2 also costimulates Tconv cells. However, intratumoral Tregs expressing more TNFR2 are able to overcome the greater resistance to suppression of intratumoral Tconv cells, resulting in a dominant immunosuppressive tumor environment.

Co-expression of TNFR2 and CD25 identifies more of the functional CD4+FOXP3+ regulatory T cells in human peripheral blood.

Previously, we found that co-expression of CD25 and TNFR2 identified the most suppressive subset of mouse Treg. In this study, we report that human peripheral blood (PB) FOXP3(+) cells present in CD25(high), CD25(low) and even CD25(-) subsets of CD4(+) cells expressed high levels of TNFR2. Consequently, TNFR2-expressing CD4(+)CD25(+) Treg included all of the FOXP3(+) cells present in the CD4(+)CD25(high) subset as well as a substantial proportion of the FOXP3(+) cells present in the CD4(+)CD25(low) subset. Flow cytometric analysis of PB identified five-fold more Treg, determined by FOXP3 expression, in the CD4(+)CD25(+)TNFR2(+) subset than in the CD4(+)CD25(high) subset. In addition, similar levels of FOXP3(+) cells were identified in both the CD4(+)CD25(+)TNFR2(+) and CD4(+)CD25(+)CD127(low/-) subsets. Furthermore, the CD4(+)CD25(+)TNFR2(+) subset expressed high levels of CTLA-4, CD45RO, CCR4 and low levels of CD45RA and CD127, a phenotype characteristic of Treg. Upon TCR stimulation, human PB CD4(+)CD25(+)TNFR2(+) cells were anergic and markedly inhibited the proliferation and cytokine production of co-cultured T-responder cells. In contrast, CD4(+)CD25(+)TNFR2(-) and CD4(+)CD25(-)TNFR2(+) T cells did not show inhibitory activity. As some non-Treg express TNFR2, the combination of CD25 and TNFR2 must be used to identify a larger population of human Treg, a population that may prove to be of diagnostic and therapeutic benefit in cancer and autoimmune diseases.

Suppression of breast cancer by chemical modulation of vulnerable zinc fingers in estrogen receptor.

Current antiestrogen therapy for breast cancer is limited by the mixed estrogenic and antiestrogenic activity of selective estrogen receptor modulators. Here we show that the function of zinc fingers in the estrogen receptor DNA-binding domain (DBD) is susceptible to chemical inhibition by electrophilic disulfide benzamide and benzisothiazolone derivatives, which selectively block binding of the estrogen receptor to its responsive element and subsequent transcription. These compounds also significantly inhibit estrogen-stimulated cell proliferation, markedly reduce tumor mass in nude mice bearing human MCF-7 breast cancer xenografts, and interfere with cell-cycle and apoptosis regulatory gene expression. Functional assays and computational analysis support a molecular mechanism whereby electrophilic agents preferentially disrupt the vulnerable C-terminal zinc finger, thus suppressing estrogen receptor-mediated breast carcinoma progression. Our results provide the proof of principle for a new strategy to inhibit breast cancer at the level of DNA binding, rather than the classical antagonism of estrogen binding.

Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma.

Myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM) contribute to cancer-related inflammation and tumor progression. While several myeloid molecules have been ascribed a regulatory function in these processes, the triggering receptors expressed on myeloid cells (TREMs) have emerged as potent modulators of the innate immune response. While various TREMs amplify inflammation, others dampen it and are emerging as important players in modulating tumor progression-for instance, soluble TREM-1 (sTREM-1), which is detected during inflammation, associates with disease progression, while TREM-2 expression is associated with tumor-promoting macrophages. We hypothesized that TREM-1 and TREM-2 might be co-expressed on tumor-infiltrating myeloid cells and that elevated sTREM-1 associates with disease outcomes, thus representing a possibility for mutual modulation in cancer. Using the 4T1 breast cancer model, we found TREM-1 and TREM-2 expression on MDSC and TAM and that sTREM-1 was elevated in tumor-bearing mice in multiple models and correlated with tumor volume. While TREM-1 engagement enhanced TNF, a TREM-2 ligand was detected on MDSC and TAM, suggesting that both TREM could be functional in the tumor setting. Similarly, we detected TREM-1 and Trem2 expression in myeloid cells in the RENCA model of renal cell carcinoma (RCC). We confirmed these findings in human disease by demonstrating the expression of TREM-1 on tumor-infiltrating myeloid cells from patients with RCC and finding that sTREM-1 was increased in patients with RCC. Finally, The Cancer Genome Atlas analysis shows that TREM1 expression in tumors correlates with poor outcomes in RCC. Taken together, our data suggest that manipulation of the TREM-1/TREM-2 balance in tumors may be a novel means to modulate tumor-infiltrating myeloid cell phenotype and function.

Histidyl-tRNA synthetase and asparaginyl-tRNA synthetase, autoantigens in myositis, activate chemokine receptors on T lymphocytes and immature dendritic cells.

Autoantibodies to histidyl-tRNA synthetase (HisRS) or to alanyl-, asparaginyl-, glycyl-, isoleucyl-, or threonyl-tRNA synthetase occur in approximately 25% of patients with polymyositis or dermatomyositis. We tested the ability of several aminoacyl-tRNA synthetases to induce leukocyte migration. HisRS induced CD4(+) and CD8(+) lymphocytes, interleukin (IL)-2-activated monocytes, and immature dendritic cells (iDCs) to migrate, but not neutrophils, mature DCs, or unstimulated monocytes. An NH(2)-terminal domain, 1-48 HisRS, was chemotactic for lymphocytes and activated monocytes, whereas a deletion mutant, HisRS-M, was inactive. HisRS selectively activated CC chemokine receptor (CCR)5-transfected HEK-293 cells, inducing migration by interacting with extracellular domain three. Furthermore, monoclonal anti-CCR5 blocked HisRS-induced chemotaxis and conversely, HisRS blocked anti-CCR5 binding. Asparaginyl-tRNA synthetase induced migration of lymphocytes, activated monocytes, iDCs, and CCR3-transfected HEK-293 cells. Seryl-tRNA synthetase induced migration of CCR3-transfected cells but not iDCs. Nonautoantigenic aspartyl-tRNA and lysyl-tRNA synthetases were not chemotactic. Thus, autoantigenic aminoacyl-tRNA synthetases, perhaps liberated from damaged muscle cells, may perpetuate the development of myositis by recruiting mononuclear cells that induce innate and adaptive immune responses. Therefore, the selection of a self-molecule as a target for an autoantibody response may be a consequence of the proinflammatory properties of the molecule itself.

Chemotactic activity of S100A7 (Psoriasin) is mediated by the receptor for advanced glycation end products and potentiates inflammation with highly homologous but functionally distinct S100A15.

Human S100A7 (psoriasin) is overexpressed in inflammatory diseases. The recently discovered, co-evolved hS100A15 is almost identical in sequence and up-regulated with hS100A7 during cutaneous inflammation. The functional role of these closely related proteins for inflammation remains undefined. By generating specific Abs, we demonstrate that hS100A7 and hS100A15 proteins are differentially expressed by specific cell types in the skin. Although highly homologous, both proteins are chemoattractants with distinct chemotactic activity for leukocyte subsets. We define RAGE (receptor for advanced glycation end products) as the hS100A7 receptor, whereas hS100A15 functions through a Gi protein-coupled receptor. hS100A7-RAGE binding, signaling, and chemotaxis are zinc-dependent in vitro, reflecting the previously reported zinc-mediated changes in the hS100A7 dimer structure. When combined, hS100A7 and hS100A15 potentiate inflammation in vivo. Thus, proinflammatory synergism in disease may be driven by the diverse biology of these almost identical proteins that have just recently evolved. The identified S100A7 interaction with RAGE may provide a novel therapeutic target for inflammation.

Regulation of the leucocyte chemoattractant receptor FPR in glioblastoma cells by cell differentiation.

The G protein-coupled formylpeptide receptor (FPR), known to mediate phagocytic leucocyte chemotaxis in response to bacterial- and host-derived agonists, was expressed by tumor cells in specimens of surgically removed more highly malignant human gliomas. In human glioblastoma cell lines, FPR activation increased cell motility, tumorigenicity and production of angiogenic factors. In studies of the mechanistic basis for the selective expression of FPR in more highly malignant gliomas, we found that the DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (Aza), while promoting the differentiation of human glioblastoma cells, downregulated FPR expression. Aza also reduced the global methylation levels in glioblastoma cells and activated the pathway of p53 tumor suppressor. Methylation-specific polymerase chain reaction revealed that Aza treatment of tumor cells reduced the methylation of p53 promoter, which was accompanied by increased expression of p53 gene and protein. In addition, overexpression of p53 in glioblastoma cells mimicked the effect of Aza treatment as shown by increased cell differentiation but reduction in FPR expression, the capacity of tumor sphere formation in soft agar and tumorigenesis in nude mice. Furthermore, Aza treatment or overexpression of the wild-type p53 in glioblastoma cells increased the binding of p53 to FPR promoter region shown by chromatin immunoprecipitation. These results indicate that increased methylation of p53 gene retains human glioblastoma cells at a more poorly differentiated phase associated with the aberrant expression of FPR as a tumor-promoting cell surface receptor.

The scaffold protein Cybr is required for cytokine-modulated trafficking of leukocytes in vivo.

Trafficking and cell adhesion are key properties of cells of the immune system. However, the molecular pathways that control these cellular behaviors are still poorly understood. Cybr is a scaffold protein highly expressed in the hematopoietic/immune system whose physiological role is still unknown. In vitro studies have shown it regulates LFA-1, a crucial molecule in lymphocyte attachment and migration. Cybr also binds cytohesin-1, a guanine nucleotide exchange factor for the ARF GTPases, which affects actin cytoskeleton remodeling during cell migration. Here we show that expression of Cybr in vivo is differentially modulated by type 1 cytokines during lymphocyte maturation. In mice, Cybr deficiency negatively affects leukocytes circulating in blood and lymphocytes present in the lymph nodes. Moreover, in a Th1-polarized mouse model, lymphocyte trafficking is impaired by loss of Cybr, and Cybr-deficient mice with aseptic peritonitis have fewer cells than controls present in the peritoneal cavity, as well as fewer leukocytes leaving the bloodstream. Mutant mice injected with Moloney murine sarcoma/leukemia virus develop significantly larger tumors than wild-type mice and have reduced lymph node enlargement, suggesting reduced cytotoxic T-lymphocyte migration. Taken together, these data support a role for Cybr in leukocyte trafficking, especially in response to proinflammatory cytokines in stress conditions.

An inhibitor of CCL2-induced chemotaxis from the fungus Leptoxyphium sp.

A biological screen used to identify inhibitors of monocyte chemotactic protein-1 (CCL2)-induced chemotaxis was applied in the activity-guided fractionation of an extract from a fungus of the genus Leptoxyphium sp. Inhibition of CCL2-induced chemotaxis was traced to a new dichlorinated diketopiperazine, cyclo(13,15-dichloro-L-Pro-L-Tyr). A structure-activity relationship (SAR) study evaluating relative activities of cyclo(13,15-dichloro-L-Pro-L-Tyr) and a nonchlorinated homologue cyclo(L-Pro-L-Tyr) showed that the dichlorinated molecule was 10- to 20-fold more active than the nonchlorinated form, while no activity was observed for cyclo(D-N-methylLeu-L-Trp).

Transactivation of the epidermal growth factor receptor by formylpeptide receptor exacerbates the malignant behavior of human glioblastoma cells.

The G protein-coupled formylpeptide receptor (FPR), which mediates leukocyte migration in response to bacterial and host-derived chemotactic peptides, promotes the chemotaxis, survival, and tumorigenesis of highly malignant human glioblastoma cells. Because glioblastoma cells may also express other receptors for growth signals, such as the epidermal growth factor (EGF) receptor (EGFR), we investigated the role of EGFR in the signaling cascade of FPR and how two receptors cross-talk to exacerbate tumor growth. We found that N-formyl-methionyl-leucyl-phenylalanine, an FPR agonist peptide, rapidly induced EGFR phosphorylation at tyrosine residue (Tyr) 992, but not residues 846, 1068, or 1173, in glioblastoma cells, whereas all these residues were phosphorylated after only EGF treatment. The FPR agonist-induced EGFR phosphorylation in tumor cells was dependent on the presence of FPR as well as Galphai proteins, and was controlled by Src tyrosine kinase. The transactivation of EGFR contributes to the biological function of FPR in glioblastoma cells because inhibition of EGFR phosphorylation significantly reduced FPR agonist-induced tumor cell chemotaxis and proliferation. Furthermore, depletion of both FPR and EGFR by short interference RNA abolished the tumorigenesis of the glioblastoma cells. Our study indicates that the glioblastoma-promoting activity of FPR is mediated in part by transactivation of EGFR and the cross-talk between two receptors exacerbates the malignant phenotype of tumor cells. Thus, targeting both receptors may yield antiglioblastoma agents superior to those targeting one of them.

Rapamycin inhibits differentiation of Th17 cells and promotes generation of FoxP3+ T regulatory cells.

Reciprocal differentiation of immunosuppressive CD4(+)CD25(+)FoxP3(+) T regulatory cells (Tregs) and proinflammatory IL-17-producing cells (Th17) from naïve CD4 cells is contingent upon the cytokine environment. Using MACS-purified CD4 cells, we found that rapamycin and cyclosporine A (CsA) potently inhibited the TGFbeta and IL-6-induced generation of IL-17-producing cells. Intriguingly, rapamycin promoted, while CsA markedly inhibited, TGFbeta-mediated generation of Tregs. The aforementioned effects of rapamycin and CsA were also observed for Flow-sorted CD4(+)CD25(-) T cells, indicating that the effect of these two immunosuppressive agents was based on their action on de novo generation of Tregs and Th17 cells from naïve CD4 cells. Our observation suggests a distinct mode of immunosuppressive action and tolerance induction by rapamycin and CsA. The capacity of rapamycin to generate immunosuppressive Tregs and to suppress differentiation of pathogenic Th17 cells furthers our understanding of the basis for the therapeutic immunosuppressive effects of rapamycin in patients with autoimmune diseases and allo-transplantation reactions.

US National Cancer Institute-China Collaborative Studies on Chinese Medicine and Cancer.

Since 2007, the US National Cancer Institute (NCI) Office of Cancer Complementary and Alternative Medicine (OCCAM), together with the Cancer Institute of the China Academy of Chinese Medical Sciences (CICACMS), institutes at China Academy of Sciences and Chinese Academy of Medical Sciences, have engaged in collaborations on Chinese medicine (CM) and cancer research. Through these collaborations, CM drugs and compounds have been studied at NCI labs. This paper summarizes the discoveries and progress on these research projects, exploring the aspects of cancer prevention, botanical drug mechanisms of action and component analysis/quality control (QC), and anticancer activity screening. These and other related projects have been presented in various jointly convened workshops and have provided the backdrop for establishing a new organization, the International Consortium for CM and Cancer, to promote international collaborations in this field.