SPARC expression induces cell cycle arrest via STAT3 signaling pathway in medulloblastoma cells

Department of Cancer Biology and Pharmacology, University of Illinois College of Medicine at Peoria, One Illini Drive, Peoria, IL-61605, United States.
Biochemical and Biophysical Research Communications (Impact Factor: 2.3). 12/2011; 417(2):874-9. DOI: 10.1016/j.bbrc.2011.12.065
Source: PubMed


Dynamic cell interaction with ECM components has profound influence in cancer progression. SPARC is a component of the ECM, impairs the proliferation of different cell types and modulates tumor cell aggressive features. We previously reported that SPARC expression significantly impairs medulloblastoma tumor growth in vivo. In this study, we demonstrate that expression of SPARC inhibits medulloblastoma cell proliferation. MTT assay indicated a dose-dependent reduction in tumor cell proliferation in adenoviral mediated expression of SPARC full length cDNA (Ad-DsRed-SP) in D425 and UW228 cells. Flow cytometric analysis showed that Ad-DsRed-SP-infected cells accumulate in the G2/M phase of cell cycle. Further, immunoblot and immunoprecipitation analyses revealed that SPARC induced G2/M cell cycle arrest was mediated through inhibition of the Cyclin-B-regulated signaling pathway involving p21 and Cdc2 expression. Additionally, expression of SPARC decreased STAT3 phosphorylation at Tyr-705; constitutively active STAT3 expression reversed SPARC induced G2/M arrest. Ad-DsRed-SP significantly inhibited the pre-established orthotopic tumor growth and tumor volume in nude-mice. Immunohistochemical analysis of tumor sections from mice treated with Ad-DsRed-SP showed decreased immunoreactivity for pSTAT3 and increased immunoreactivity for p21 compared to tumor section from mice treated with mock and Ad-DsRed. Taken together our studies further reveal that STAT3 plays a key role in SPARC induced G2/M arrest in medulloblastoma cells. These new findings provide a molecular basis for the mechanistic understanding of the effects of SPARC on medulloblastoma tumor cell proliferation.

Download full-text


Available from: Ganji Purnachandra Nagaraju, Jun 10, 2014
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Natural Killer cells (NK) are critical components of the innate immune system. Often referred to by their morphology, these large granular lymphocytes (LGLs) are bone marrow-derived lymphocytes and can be found throughout the body. NK cells reside in the liver, lymph nodes, spleen, thymus, and mucosal-associated lymphoid tissues (MALT). Importantly, NK cells also circulate throughout the blood where they function as surveyors of the body and are armed to eliminate malignant, infected, damaged, or foreign cells. NK cells function by a dual receptor system. That is, NK receptors are broadly categorized as inhibitory or activating. It is a fine balance, or lack thereof, that dictates the function of an NK cell. Unlike their T and B cell adaptive counterparts, NK cell receptors (NKR) are germline encoded and do not undergo gene rearrangement. NKRs are expressed in a variegated but overlapping fashion such that different cell subsets in the NK compartment elaborate different combinations of activating and inhibitory NKR. Varying the array of NKRs used by each subset increases the potential specificities of the NK compartment, while retaining tolerance to self. Thus, a diverse and balanced NK cell receptor repertoire (NKRR) is extremely important in order for this lineage to respond to various immunologic challenges and to do so in a normal, effective manner. As we have previously shown, aberrations in the expression of NKRs or downstream signaling can lead to severe immune deficiency, as observed in SHIP-deficient mice. We also showed that in the absence of SHIP-1, 2B4 becomes highly upregulated, functioning as a dominant inhibitory receptor and rendering the SHIP-1-deficient NK cell unresponsive to complex tumor targets. Traditionally MHC-I inhibitory ligands are largely responsible for the regulation of NK function. However, we show here that 2B4, which mediates MHC-I-independent inhibition, is required for formation of a normal NKRR, NK homeostasis, and effector functions. Moreover, in the absence of 2B4 and SHIP-1, NK cells have improper licensing, or education. In addition to SHIP-1 and 2B4 we show that the nature of the MHC-I ligands also play a significant role in repertoire formation, NK effector functions, and NK cell education. As described above, NK cells are critical components of the immune system. Understanding how NK cell biology and function are regulated, or affected in the context of pathology is of high significance. NK function is often severely impaired in a diseased state, and more importantly, NK cells are frequently adversely affected by the treatments themselves. Here we sought out to determine the effects of an immunomodulating drug, lenalidomide, on the biology and function of healthy NK cells. Lenalidomide is a unique drug that displays immune enhancing functions yet can be cytotoxic to tumor cells. However, lenalidomide treatment can result in immune suppression and severe cytopenia, and has the ability to impair NK viability. We show here that if used in combination with cytokine treatment (e.g. IL-2 or IL-15), many of these negative affects can be overcome. Furthermore, we show that lenalidomide treatment results in what appears to be an NK activating phenotype with a down-modulation of inhibitory KIRs and upregulation of CD16. Lenalidomide also leads to a sustained and robust activation of STAT5 and consequential increase in perforin and granzyme B. Finally, we find that treatment with lenalidomide in combination with IL-2 or IL-15 enhances the expression of IL-Rβ and IL-2Rγ chains, a presumed mechanism of action, which may provide a positive feedback loop. These findings have important clinical application. We propose that using lenalidomide in combination with IL-15 can augment its immune activating effects, while minimizing unwanted cytopenias.
    Preview · Article ·
  • [Show abstract] [Hide abstract]
    ABSTRACT: Mitotically inactivated feeder cells such as mouse embryonic fibroblast (MEFs) cells have been widely applied for physical and physiological support in the pluripotency maintenance of human pluripotent stem cells (hPSCs). However, accurate supporting mechanism or factors of feeder cells are poorly understood. Here, we isolated differentially expressed genes between wild-type MEFs and mitotically inactivated MEFs (miMEFs) by employing annealing control primer-based GeneFishing polymerase chain reaction. We identified a secreted protein acidic cysteine-rich glycoprotein (SPARC) gene that is upregulated in miMEFs. Suppression of SPARC expression in miMEFs using small interference RNA (siRNA) displayed gradual detachment of miMEFs. Furthermore, we found a significant reduction of OCT4- and SSEA3-positive hPS cell population maintained on SPARC siRNA-miMEFs compared to on miMEFs by flow cytometrical analysis. These findings suggest that SPARC plays a critical role in the maintenance of miMEFs without loss of cell number and might be a key component for supporting the culture of hPSCs.
    No preview · Article · May 2013 · In Vitro Cellular & Developmental Biology - Animal
  • [Show abstract] [Hide abstract]
    ABSTRACT: Despite advances in the treatment of diabetic nephropathy (DN), currently available therapies have not prevented the epidemic of progressive chronic kidney disease (CKD). The morbidity of CKD, and the inexorable increase in the prevalence of end stage renal disease, demands more effective approaches to prevent and treat progressive CKD. We undertook next generation sequencing in a rat model of diabetic nephropathy to study in depth the pathogenic alterations involved in DN with progressive CKD. We employed the obese, diabetic ZS rat, a model that develops diabetic nephropathy, characterized by progressive CKD, inflammation and fibrosis, the hallmarks of human disease. We then used RNA-seq to examine the combined effects of renal cells and infiltrating inflammatory cells acting as a pathophysiological unit. The comprehensive systems biology analysis of progressive CKD revealed multiple interactions of altered genes that were integrated into morbid networks. These pathological gene assemblies lead to renal inflammation, promote apoptosis and cell cycle arrest in progressive CKD. Moreover, in what is clearly a major therapeutic challenge, multiple and redundant pathways were found to be linked to renal fibrosis, a major cause of kidney loss. We conclude that systems biology applied to progressive CKD in DN can be used to develop novel therapeutic strategies directed to restore critical anomalies in affected gene networks.
    No preview · Article · Jun 2013 · Physiological Genomics
Show more