Proteomic profiling of human retinal pigment epithelium exposed to an advanced glycation-modified substrate

PURPOSE: The retinal pigment epithelium (RPE) and underlying Bruch’s membrane undergo significant modulation during ageing. Progressive, age-related modifications of lipids and proteins by advanced glycation end products (AGEs) at this cell–substrate interface have been implicated in RPE dysfunction and the progression to age-related macular degeneration (AMD). The pathogenic nature of these adducts in Bruch’s membrane and their influence on the overlying RPE remains unclear. This study aimed to identify alterations in RPE protein expression in cells exposed to AGE-modified basement membrane (AGE-BM), to determine how this “aged” substrate impacts RPE function and to map the localisation of identified proteins in ageing retina. METHODS: Confluent ARPE-19 monolayers were cultured on AGE-BM and native, non-modified BM (BM). Following 28-day incubation, the proteome was profiled using 2-dimensional gel electrophoresis (2D), densitometry and image analysis was employed to map proteins of interest that were identified by electrospray ionisation mass spectrometry (ESI MS/MS). Immunocytochemistry was employed to localise identified proteins in ARPE-19 monolayers cultured on unmodified and AGE-BM and to analyze aged human retina. RESULTS: Image analysis detected altered protein spot densities between treatment groups, and proteins of interest were identified by LC ESI MS/MS which included heat-shock proteins, cytoskeletal and metabolic regulators. Immunocytochemistry revealed deubiquitinating enzyme ubiquitin carboxyterminal hydrolase-1 (UCH-L1), which was upregulated in AGE-exposed RPE and was also localised to RPE in human retinal sections. CONCLUSIONS: This study has demonstrated that AGE-modification of basement membrane alters the RPE proteome. Many proteins are changed in this ageing model, including UCHL-1, which could impact upon RPE degradative capacity. Accumulation of AGEs at Bruch”s membrane could play a significant role in age-related dysfunction of the RPE

High-risk human papillomaviruses and epstein-barr virus presence and crosstalk in human oral carcinogenesis

Oral cancer is the most frequent type of malignancy of the head and neck area worldwide. This cancer is a major cause of mortality in several parts of the world especially in certain Asian countries. In fact, the majority of cancer deaths are the result of metastasis, either directly due to tumor involvement of critical organs or indirectly due to therapeutic side effects. On the other hand, it is estimated that 10–20% of human cancers are linked to virus infections including Epstein-Barr virus (EBV) and high-risk human papillomaviruses (HPVs) especially types 16, 18, and 33, which cumulatively infect 80–90% of the population worldwide. Moreover, it has been reported that oncoproteins of high-risk HPV type 16 can covert noninvasive and nonmetastatic human cancer cells into invasive and metastatic form. It was pointed out that high-risk HPVs and EBV are important etiological factors in human oral cancer, since around 35 and 55% of these cancers are positive for these viruses, respectively. In addition, it was recently revealed that high-risk HPVs and EBV can be copresent in human oral cancer, and their copresence is associated with high-grade invasive carcinomas. Thus, it is evident that high-risk HPVs and EBV oncoproteins play an important role in the initiation and progression of human oral cancer. In this chapter, we will overview the presence and role of HPVs and EBV in this frequent malignancy, more specifically; we will focus on the role of their oncoproteins and their interactions with other oncogenes in human oral carcinogenesis.Scopu