BackgroundRetinoblastoma (Rb) is the most common primary intraocular tumor in children. Local treatment of the intraocular disease is usually effective if diagnosed early; however advanced Rb can metastasize through routes that involve invasion of the choroid, sclera and optic nerve or more broadly via the ocular vasculature. Metastatic Rb patients have very high mortality rates. While current therapy for Rb is directed toward blocking tumor cell division and tumor growth, there are no specific treatments targeted to block Rb metastasis. Two such targets are matrix metalloproteinases-2 and -9 (MMP-2, −9), which degrade extracellular matrix as a prerequisite for cellular invasion and have been shown to be involved in other types of cancer metastasis. Cancer Clinical Trials with an anti-MMP-9 therapeutic antibody were recently initiated, prompting us to investigate the role of MMP-2, −9 in Rb metastasis.MethodsWe compare MMP-2, −9 activity in two well-studied Rb cell lines: Y79, which exhibits high metastatic potential and Weri-1, which has low metastatic potential. The effects of inhibitors of MMP-2 (ARP100) and MMP-9 (AG-L-66085) on migration, angiogenesis, and production of immunomodulatory cytokines were determined in both cell lines using qPCR, and ELISA. Cellular migration and potential for invasion were evaluated by the classic wound-healing assay and a Boyden Chamber assay.ResultsOur results showed that both inhibitors had differential effects on the two cell lines, significantly reducing migration in the metastatic Y79 cell line and greatly affecting the viability of Weri-1 cells. The MMP-9 inhibitor (MMP9I) AG-L-66085, diminished the Y79 angiogenic response. In Weri-1 cells, VEGF was significantly reduced and cell viability was decreased by both MMP-2 and MMP-9 inhibitors. Furthermore, inhibition of MMP-2 significantly reduced secretion of TGF-β1 in both Rb models.ConclusionsCollectively, our data indicates MMP-2 and MMP-9 drive metastatic pathways, including migration, viability and secretion of angiogenic factors in Rb cells. These two subtypes of matrix metalloproteinases represent new potential candidates for targeted anti-metastatic therapy for Rb.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-017-3418-y) contains supplementary material, which is available to authorized users.

Whole organ culture of the spiral ganglion region is a resourceful model system facilitating manipulation and analysis of live sprial ganglion neurons (SGNs). Three-dimensional (3D) cultures have been demonstrated to have many biomedical applications, but the effect of 3D culture in maintaining the SGNs structure and function in explant culture remains uninvestigated. In this study, we used the matrigel to encapsulate the spiral ganglion region isolated from neonatal mice. First, we optimized the matrigel concentration for the 3D culture system and found the 3D culture system protected the SGNs against apoptosis, preserved the structure of spiral ganglion region, and promoted the sprouting and outgrowth of SGNs neurites. Next, we found the 3D culture system promoted growth cone growth as evidenced by a higher average number and a longer average length of filopodia and a larger growth cone area. 3D culture system also significantly elevated the synapse density of SGNs. Last, we found that the 3D culture system combined with neurotrophic factors had accumulated effects in promoting the neurites outgrowth compared with 3D culture or NFs treatment only groups. Together, we conclude that the 3D culture system preserves the structure and function of SGN in explant culture.

BackgroundType 2 diabetes is commonly characterized by insulin deficiency and decreased sensitivity of insulin receptors, leading to a chronic state of hyperglycemia in individuals. Disease progression induces changes in the immune profile that engenders a chronic inflammatory condition. Thiazolidinedione (TDZ) drugs, such as Pioglitazone (Pio), aid in controlling disease symptoms. While the mechanisms by which Pio controls hyperglycemia are beginning to be understood, relatively little is known about the effects of Pio on suppression of the systemic immune phenotype, attributed to visceral adipose tissue and macrophages.MethodsHere, we utilize the recently developed BBDZR/Wor type 2 diabetes rat model to test our hypothesis that a selective in vivo growth of CD3+T cells in the spleen contributes to the increase in T lymphocytes, including Tregs, independent of visceral adipose tissue. We investigated the systemic effects of Pio on multifactorial aspects of the disease-induced immune phenotype both in vivo and in vitro in normal, non-diabetic animals and in disease.ResultsOur work revealed that Pio reversed the lymphopenic status of diabetic rats, in part by an increase in CD3+ T lymphocytes and related subsets. Moreover, we found evidence that Pio caused a selective growth of newly differentiated T lymphocytes, based on the presence of recent thymic emigrants in vivo. To investigate effects of Pio on the inflammatory milieu, we examined the production of the signature cytokines TNF-α and IL-1β and found they were reduced by Pio-treatment, while the levels of IL-4, an anti-inflammatory mediator, were significantly increased in a Pio-dependent manner. The increase in IL-4 production, although historically attributed to macrophages from visceral adipose tissue under other conditions, came also from CD3+ T lymphocytes from the spleen, suggesting splenocytes contribute to the Pio-induced shift towards an anti-inflammatory phenotype.ConclusionsWe show for the first time that Pio treatment significantly suppresses the systemic inflammatory status in the BBDZR/Wor type 2 diabetes rat model by the selective growth of newly differentiated CD3+ T cells and by increasing CD3+IL-4 production in immigrant spleen lymphocytes.

Cite this articleGoldsmith ZK, McEwen MW, Kibe MW, et al. Sirtuin inhibitor as a novel cell cycle checkpoint and regulator of the TP53-MDM2 pathway in uveal melanoma. ABSTRACT PurposeThe liver is the most common site of uveal melanoma (UM) metastasis with approximately 50% of UM patients being affected. With no proven therapies that mitigate metastases the mortality rate is 85% within the first year after detection of the liver disease. In this study, we provide a mechanistic understanding of the de-regulation of the TP53-MDM2 pathway in UM, which plays a central role in tumor biology. MethodsWe investigated the TP53-MDM2 signaling pathway in the microenvironment of liver metastases taken from both a murine orthotopic xenograft and post-mortem metastatic UM human liver. These findings were studied in-depth using both primary and metastatic UM cell lines treated with the MDM2 antagonist Nutlin-3a and the sirtuin inhibitor and transcriptional activator of TP53, Tenovin-6. ResultsDe-regulation of the TP53-MDM2 signaling pathway is specific to the liver microenvironment, providing a survival mechanism for UM metastases. Tenovin-6, not Nutlin-3a, reduced UM cell survival by increasing the percentage of cell death and reducing the percentage of proliferating cells. Tenovin-6 increased acetylation of p53, reduced ubiquitination of the protein, and acted as a cell cycle regulator. ConclusionOur findings suggest that in patients with metastatic UM de-regulation of TP53-MDM2 signaling pathway promotes growth of the liver metastases and provides pre-clinical information on the potential of targeting of the TP53-MDM2 signaling pathway via Tenovin-6.