Large-scale analysis of protein expression changes in human keratinocytes immortalized by human papilloma virus type 16 E6 and E7 oncogenes

<p>Abstract</p> <p>Background</p> <p>Infection with high-risk type human papilloma viruses (HPVs) is associated with cervical carcinomas and with a subset of head and neck squamous cell carcinomas. Viral E6 and E7 oncogenes cooperate to achieve cell immortalization by a mechanism that is not yet fully understood. Here, human keratinocytes were immortalized by long-term expression of HPV type 16 E6 or E7 oncoproteins, or both. Proteomic profiling was used to compare expression levels for 741 discrete protein features.</p> <p>Results</p> <p>Six replicate measurements were performed for each group using two-dimensional difference gel electrophoresis (2D-DIGE). The median within-group coefficient of variation was 19–21%. Significance of between-group differences was tested based on Significance Analysis of Microarray and fold change. Expression of 170 (23%) of the protein features changed significantly in immortalized cells compared to primary keratinocytes. Most of these changes were qualitatively similar in cells immortalized by E6, E7, or E6/7 expression, indicating convergence on a common phenotype, but fifteen proteins (~2%) were outliers in this regulatory pattern. Ten demonstrated opposite regulation in E6- and E7-expressing cells, including the cell cycle regulator p16^INK4a; the carbohydrate binding protein Galectin-7; two differentially migrating forms of the intermediate filament protein Cytokeratin-7; HSPA1A (Hsp70-1); and five unidentified proteins. Five others had a pattern of expression that suggested cooperativity between the co-expressed oncoproteins. Two of these were identified as forms of the small heat shock protein HSPB1 (Hsp27).</p> <p>Conclusion</p> <p>This large-scale analysis provides a framework for understanding the cooperation between E6 and E7 oncoproteins in HPV-driven carcinogenesis.</p

Reduced vasorin enhances angiotensin II signaling within the aging arterial wall

The glycosylated protein vasorin physically interacts with the transforming growth factor-beta1 (TGF-β1) and functionally attenuates its fibrogenic signaling in the vascular smooth muscle cells (VSMCs) of the arterial wall. Angiotensin II (Ang II) amplifies TGF-β1 activation in the VSMCs of the arterial wall with aging. In this study, we hypothesized that a reduced expression of the protein vasorin plays a contributory role in magnifying Ang II-associated fibrogenic signaling in the VSMCs of the arterial wall with aging. The current study shows that vasorin mRNA and protein expression were significantly decreased both in aortic wall and VSMCs from old (30 mo) vs. young (8 mo) FXBN rats. Exposing young VSMCs to Ang II reduced vasorin protein expression to the levels of old untreated cells while treating old VSMCs with the Ang II type AT1 receptor antagonist Losartan upregulated vasorin protein expression up to the levels of young. The physical interaction between vasorin and TGF-β1 was significantly decreased in old vs. young VSMCs. Further, exposing young VSMCs to Ang II increased the levels of matrix metalloproteinase type II (MMP-2) activation and TGF-β1 downstream molecules p-SMAD-2/3 and collagen type I production up to the levels of old untreated VSMCs, and these effects were substantially inhibited by overexpressing vasorin. Administration of Ang II to young rats (8 mo) for 28 days via an osmotic minipump markedly reduced the expression of vasorin. Importantly, vasorin protein was effectively cleaved by activated MMP-2 both in vitro and in vivo. Administration of the MMP inhibitor, PD 166793, for 6 mo to young adult (18 mo) via a daily gavage markedly increased levels of vasorin in the aortic wall. Thus, reduced vasorin amplifies Ang II profibrotic signaling via an activation of MMP-2 in VSMCs within the aging arterial wall

Foot-and-Mouth Disease Virus Serotype A in Egypt

We describe the characterization of a foot-and-mouth disease (FMD) serotype A virus responsible for recent outbreaks of disease in Egypt. Phylogenetic analysis of VP1 nucleotide sequences demonstrated a close relationship to recent FMD virus isolates from East Africa, rather than to viruses currently circulating in the Middle East

Evaluation Research and Institutional Pressures: Challenges in Public-Nonprofit Contracting

This article examines the connection between program evaluation research and decision-making by public managers. Drawing on neo-institutional theory, a framework is presented for diagnosing the pressures and conditions that lead alternatively toward or away the rational use of evaluation research. Three cases of public-nonprofit contracting for the delivery of major programs are presented to clarify the way coercive, mimetic, and normative pressures interfere with a sound connection being made between research and implementation. The article concludes by considering how public managers can respond to the isomorphic pressures in their environment that make it hard to act on data relating to program performance.This publication is Hauser Center Working Paper No. 23. The Hauser Center Working Paper Series was launched during the summer of 2000. The Series enables the Hauser Center to share with a broad audience important works-in-progress written by Hauser Center scholars and researchers

The Society for Immunotherapy of Cancer consensus statement on immunotherapy for the treatment of squamous cell carcinoma of the head and neck (HNSCC).

Head and neck cancers, including those of the lip and oral cavity, nasal cavity, paranasal sinuses, oropharynx, larynx and nasopharynx represent nearly 700,000 new cases and 380,000 deaths worldwide per annum, and account for over 10,000 annual deaths in the United States alone. Improvement in outcomes are needed for patients with recurrent and or metastatic squamous cell carcinoma of the head and neck (HNSCC). In 2016, the US Food and Drug Administration (FDA) granted the first immunotherapeutic approvals - the anti-PD-1 immune checkpoint inhibitors nivolumab and pembrolizumab - for the treatment of patients with recurrent squamous cell carcinoma of the head and neck (HNSCC) that is refractory to platinum-based regimens. The European Commission followed in 2017 with approval of nivolumab for treatment of the same patient population, and shortly thereafter with approval of pembrolizumab monotherapy for the treatment of recurrent or metastatic HNSCC in adults whose tumors express PD-L1 with a ≥ 50% tumor proportion score and have progressed on or after platinum-containing chemotherapy. Then in 2019, the FDA granted approval for PD-1 inhibition as first-line treatment for patients with metastatic or unresectable, recurrent HNSCC, approving pembrolizumab in combination with platinum and fluorouracil for all patients with HNSCC and pembrolizumab as a single agent for patients with HNSCC whose tumors express a PD-L1 combined positive score ≥ 1. These approvals marked the first new therapies for these patients since 2006, as well as the first immunotherapeutic approvals in this disease. In light of the introduction of these novel therapies for the treatment of patients with head and neck cancer, The Society for Immunotherapy of Cancer (SITC) formed an expert committee tasked with generating consensus recommendations for emerging immunotherapies, including appropriate patient selection, therapy sequence, response monitoring, adverse event management, and biomarker testing. These consensus guidelines serve as a foundation to assist clinicians\u27 understanding of the role of immunotherapies in this disease setting, and to standardize utilization across the field for patient benefit. Due to country-specific variances in approvals, availability and regulations regarding the discussed agents, this panel focused solely on FDA-approved drugs for the treatment of patients in the U.S

Differential recognition of HIV-stimulated IL-1? and IL-18 secretion through NLR and NAIP signalling in monocyte-derived macrophages

Macrophages are important drivers of pathogenesis and progression to AIDS in HIV infection. The virus in the later phases of the infection is often predominantly macrophage-tropic and this tropism contributes to a chronic inflammatory and immune activation state that is observed in HIV patients. Pattern recognition receptors of the innate immune system are the key molecules that recognise HIV and mount the inflammatory responses in macrophages. The innate immune response against HIV-1 is potent and elicits caspase-1-dependent pro-inflammatory cytokine production of IL-1β and IL-18. Although, NLRP3 has been reported as an inflammasome sensor dictating this response little is known about the pattern recognition receptors that trigger the “priming” signal for inflammasome activation, the NLRs involved or the HIV components that trigger the response. Using a combination of siRNA knockdowns in monocyte derived macrophages (MDMs) of different TLRs and NLRs as well as chemical inhibition, it was demonstrated that HIV Vpu could trigger inflammasome activation via TLR4/NLRP3 leading to IL-1β/IL-18 secretion. The priming signal is triggered via TLR4, whereas the activation signal is triggered by direct effects on Kv1.3 channels, causing K+ efflux. In contrast, HIV gp41 could trigger IL-18 production via NAIP/NLRC4, independently of priming, as a one-step inflammasome activation. NAIP binds directly to the cytoplasmic tail of HIV envelope protein gp41 and represents the first non-bacterial ligand for the NAIP/NLRC4 inflammasome. These divergent pathways represent novel targets to resolve specific inflammatory pathologies associated with HIV-1 infection in macrophages