Programmed cell death 4 (Pdcd4) is a novel repressor of in vitro transformation. Pdcd4 directly inhibits the helicase activity of eukaryotic translation initiation factor 4A, a component of the translation initiation complex. To ascertain whether Pdcd4 suppresses tumor development in vivo, we have generated transgenic mice that overexpress Pdcd4 in the epidermis (K14-Pdcd4). K14-regulated Pdcd4 expression caused a neonatal short-hair phenotype due to early catagen entry compared with matched wild-type siblings. In response to the 7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA) mouse skin carcinogenesis protocol, K14-Pdcd4 mice showed significant reductions in papilloma formation, carcinoma incidence, and papilloma-to-carcinoma conversion frequency compared with wild-type mice. The translational efficiency of an mRNA engineered to form a structured 5V untranslated region (UTR) was attenuated in primary keratinocytes when Pdcd4 was overexpressed. Pdcd4 inhibited by 46% TPA-induced activator protein-1 (AP-1)-dependent transcription, an event required for tumorigenesis. CDK4 and ornithine decarboxylase (ODC) are candidates for Pdcd4-regulated translation as their mRNAs contain 5Vstructured UTRs. In K14-Pdcd4 primary keratinocytes expressing activated Ha-Ras to mimic DMBA-initiated epidermis, ODC and CDK4 protein levels were decreased by 40% and 46%, respectively. Expression of a protein encoded by 5V unstructured mRNA showed no change. These results extend to an in vivo model the observations that Pdcd4 inhibits both translation initiation and AP-1 activation while decreasing benign tumor development and malignant progression. The K14-Pdcd4 mice seem to validate translation initiation as a novel target for cancer prevention. (Cancer Res 2005; 65(14): 6034-41)

Bone is a dynamic tissue that undergoes renewal throughout life by a process whereby osteoclasts resorb worn bone and osteoblasts synthesize new bone. Imbalances in bone turnover lead to bone loss and development of osteoporosis and ultimately fracture, a debilitating condition with high morbidity and mortality. Silica is a ubiquitous biocontaminant that is considered to have high biocompatibility. We report that silica nanoparticles mediate potent inhibitory effects on osteoclasts and stimulatory effects on osteoblasts in vitro. The mechanism of bioactivity is a consequence of an intrinsic capacity to antagonize activation of NF-κB, a signal transduction pathway required for osteoclastic bone resorption, but inhibitory to osteoblastic bone formation. We further demonstrate that silica nanoparticles promote a significant enhancement of bone mineral density (BMD) in mice in vivo providing a proof of principle for the potential application of silica nanoparticles as a pharmacological agent to enhance BMD and protect against bone fracture.

Many key processes central to bone formation and homeostasis require the involvement of osteoblasts, cells responsible for accumulation and mineralization of the extracellular matrix (ECM). During this complex and only partially understood process, osteoblasts generate and secrete matrix vesicles (MVs) into the ECM to initiate mineralization. Although they are considered an important component of mineralization process, MVs still remain a mystery. To better understand their function and biogenesis, a proteomic analysis of MVs has been conducted. MVs were harvested by two sample preparation approaches and mass spectrometry was utilized for protein identification. A total of 133 proteins were identified in common from the two MV preparations, among which were previously known proteins, such as annexins and peptidases, along with many novel proteins including a variety of enzymes, osteoblast-specific factors, ion channels, and signal transduction molecules, such as 14-3-3 family members and Rab-related proteins. To compare the proteome of MV with that of the ECM we conducted a large-scale proteomic analysis of collagenase digested mineralizing osteoblast matrix. This analysis resulted in the identification of 1,327 unique proteins. A comparison of the proteins identified from the two MV preparations with the ECM analysis revealed 83 unique, non-redundant proteins identified in all three samples. This investigation represents the first systematic proteomic analysis of MVs and provides insights into both the function and origin of these important mineralization-regulating vesicles.

A new synthetic method has been developed to prepare fluorescent silica nanoparticles without employing isothiocyanated dye molecules and (3-aminopropyl)triethoxysilane (APS) for the thiourea linkage formation; the resulting fluorescent silica nanoparticles show excellent photochemical, thermal and pH stabilities and a good biocompatibility with over 85% viability from various cell types.Since the preparation method of silica nanoparticles was developed by Werner Stöber, 1 many researchers have focused on attempts to control the size and uniformity, as well as to utilize them to various fields including biological application. Silica has several advantages including its ease of preparation through the hydrolysis-condensation reaction from relatively inexpensive precursor molecules such as tetraethyl orthosilicate (TEOS) in the presence of acid-or base-catalysts, the possibility of surface modification with various well-studied organosilicon compounds,2 and its non-acute toxicity.3a,b As the surface silanol groups can be easily modified by various functional groups and modified silica nanoparticles can effectively penetrate the cell membrane, there has been a great amount of research effort to use them as carriers for the delivery of drugs or genes.3a,c,d , 4 To improve the application of silica nanoparticles to biological research, fluorescent dye molecules were introduced into silica nanoparticles by using a thiourea-linkage forming reaction through amino-terminated alkyltrialkoxysilane compounds such as (3-aminopropyl)triethoxysilane (APS) and dye molecules having an isothiocyanate functional group, i.e. Rhodamine isothiocyanate (RITC) and Fluorescein isothiocyanate (FITC).5 This photostable fluorescent property has become one of the most appreciated functionalities of fluorescent silica nanoparticles and has opened a new era of bioimaging.6 -8a Although this thiourea-linkage formation reaction is simple and well studied as a useful bioconjugation method,9 it has several drawbacks in preparing fluorescent silica nanoparticles. First, an excess amount of APS is usually used in order to incorporate all the possible dye molecules into silica nanoparticles (about two times excess compared to the amount of isothiocyanated dye molecule such as RITC or FITC), and it is cocondensed with TEOS during the silica formation process. Although the amino terminal groups on the nanoparticle surface could be used for the further chemical linkage reactions such as conjugation with drugs or specific binding antibodies,8b,d the amount of amino terminal groups was not reproducibly controllable and the excess amount of APS sometimes adversely NIH Public Access Author ManuscriptChem Commun (Camb). Author manuscript; available in PMC 2010 June 1. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript affected the size uniformity.10a Second, it is reported that primary amine groups on the surface make silica nanoparticles stable and dispersible only in acidic condition, but lead to precipitation in neutral ...