Tissue inhibitor of metalloproteinase 1 is preferentially expressed in Th1 and Th17 T-helper cell subsets and is a direct STAT target gene.

CD4(+) T helper (Th) cells differentiate into distinct effector subsets that are critical for host defense, but are also implicated in the pathogenesis of autoimmune disorders. Thelper17 (Th17) cells in particular are emerging as important drivers of multiple diseases including psoriasis, spondyloarthropathy and multiple sclerosis. To gain insight into the function of Th17 cells, we performed transcriptional profiling in hopes of elucidating products not previously recognized as being functionally relevant in these T cells. Herein, we demonstrate that tissue inhibitor of metalloproteinase 1 (TIMP1), a secreted protein with pleiotropic effects on cellular growth, survival and integrity of the extracellular matrix, is preferentially produced by Th17 and Th1 cells. We further show that Th1 and Th17 cell TIMP1 regulation follows separate mechanisms with a requirement for STAT4 in the former and STAT3 in the latter. Finally, we demonstrate that when restricted to T cells, expression of TIMP1 promotes neuropathology in experimental allergic encephalomyelitis.

Endogenous angiogenesis inhibitor blocks tumor growth via direct and indirect effects on tumor microenvironment.

Tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) belongs to a small family of endogenous proteins that inhibits a group of enzymes, the matrix metalloproteinases (MMPs). TIMP-2 inhibits endothelial cell proliferation and migration in vitro and angiogenesis in vivo, through MMP-dependent and -independent mechanisms. However, little is known regarding the contribution of these mechanisms to the antitumor effects of TIMP-2. Using a retroviral delivery system, we stably overexpressed TIMP-2 and its mutant Ala+TIMP-2 (devoid of MMP inhibitory activity) in human adenocarcinoma A549 cells. Using real time PCR, and enzyme-linked immunosorbent assay (ELISA), we confirmed enhanced TIMP-2 expression and its MMP inhibitory activity by reverse zymography. In vitro, growth assays suggested that TIMP-2 and Ala+TIMP-2 did not alter basal cell proliferation rates, however, tumor cell migration and invasion were inhibited. In vivo, both TIMP-2 and Ala+TIMP-2 A549 xenografts exhibited reduced growth rate, CD31 immunostaining indicated decreased intratumoral microvascular density, and TUNEL demonstrated enhanced tumor cell apoptosis. Immunoblotting and immunohistochemical analyses of A549 xenograft tissues with either phospho-FAK (Tyr397) or phospho-AKT (Ser473) showed decreased activation in both TIMP-2 and Ala+TIMP-2 tumor cells. We conclude that TIMP-2-mediated inhibition of tumor growth occurs, at least in part, independently of MMP inhibition, and is a consequence of both direct effects of TIMP-2 on tumor cells and modulation of the tumor microenvironment.

Synthesis of novel cyclic NGR/RGD peptide analogs via on resin click chemistry.

Targeted drug deliveries as well as high resolution imaging of cancerous tissues and organs via specific cancer cell markers have become important in chemotherapeutic interventions of cancer treatment. Short peptides such as RGD and NGR are showing promising results for targeted drug delivery and in vivo imaging. We have applied on resin Huisgen's 1,3-dipolar cycloaddition to synthesize new cyclic RGD and NGR peptide analogs. Preliminary binding assays of these new analogs by fluorescence polarization indicates specific binding to purified CD13 (Aminopeptidase N) and cell lysates from MCF-7 and SKOV-3 cancer cell lines.

Calcineurin activation influences muscle phenotype in a muscle-specific fashion.

BACKGROUND: The calcium activated protein phosphatase 2B, also known as calcineurin, has been implicated as a cell signaling molecule involved with transduction of physiological signals (free cytosolic Ca2+) into molecular signals that influence the expression of phenotype-specific genes in skeletal muscle. In the present study we address the role of calcineurin in mediating adaptations in myosin heavy chain (MHC) isoform expression and muscle mass using 3-month old wild-type (WT) and transgenic mice displaying high-level expression of a constitutively active form of calcineurin (MCK-CN* mice). RESULTS: Slow muscles, e.g., soleus, were significantly larger (by ~24%), whereas fast muscles, e.g., medial gastrocnemius (MG) and tibialis anterior were significantly smaller (by ~26 and ~16%, respectively) in MCK-CN* mice compared to WT. The masses of mixed phenotype muscles, such as the plantaris and the extensor digitorum longus, were not significantly changed from WT. The soleus, plantaris, MG and diaphragm displayed shifts toward slower MHC isoforms, e.g., soleus from WT mice contained ~52% MHC-I, ~39% MHC-IIa, and ~9% MHC-IIx, whereas MCK-CN* mice had ~67% MHC-I, ~26% MHC-IIa, and ~7% MHC-IIx. The specific isoforms that were either up or down-regulated were muscle-specific. For instance, the proportion of MHC-IIa was decreased in the soleus and diaphragm, but increased in the plantaris and MG of MCK-CN* mice. Also, the proportion of MHC-IIx was unchanged in the soleus, decreased in the diaphragm and increased in the plantaris and MG of MCK-CN* relative to WT mice. Fast to slow shifts in fiber type proportions were evident for the plantaris, but not the soleus. Fast, but not slow, plantaris fibers of MCK-CN* mice had higher oxidative and lower glycolytic properties than WT. CONCLUSION: These data suggest that calcineurin activation can influence muscle phenotype and that the specific influence of calcineurin activation on the phenotypic and mass characteristics of a muscle is dependent upon the original phenotypic state of the muscle.