Promoter activity of Merkel cell Polyomavirus variants in human dermal fibroblasts and a Merkel cell carcinoma cell line.

BACKGROUND: Merkel cell polyomavirus (MCPyV) is a human polyomavirus that establishes a life-long harmless infection in most individuals, with dermal fibroblasts believed to be the natural host cell. However, this virus is the major cause of Merkel cell carcinoma (MCC), an aggressive skin cancer. Several MCPyV variants with polymorphism in their promoter region have been isolated, but it is not known whether these differences affect the biological properties of the virus. METHODS: Using transient transfection studies in human dermal fibroblasts and the MCC cell line MCC13, we compared the transcription activity of the early and late promoters of the most commonly described non-coding control region MCPyV variant and six other isolates containing specific mutation patterns. RESULTS: Both the early and late promoters were significantly stronger in human dermal fibroblasts compared with MCC13 cells, and a different promoter strength between the MCPyV variants was observed. The expression of full-length large T-antigen, a viral protein that regulates early and late promoter activity, inhibited early and late promoter activities in both cell lines. Nonetheless, a truncated large T-antigen, which is expressed in virus-positive MCCs, stimulated the activity of its cognate promoter. CONCLUSION: The promoter activities of all MCPyV variants tested was stronger in human dermal fibroblasts, a cell line that supports viral replication, than in MCC13 cells, which are not permissive for MCPyV. Truncated large T-antigen, but not full-length large T-antigen stimulated viral promoter activity. Whether, the difference in promoter strength and regulation by large T-antigen may affect the replication and tumorigenic properties of the virus remains to be determined.

Whole-genome sequencing of ocular Chlamydia trachomatis isolates from Gadarif State, Sudan.

BACKGROUND: Trachoma, caused by ocular Chlamydia trachomatis, is the leading infectious cause of blindness worldwide. Sudan first reported trachoma in the 1930s and has since been consistently endemic. Ocular C. trachomatis previously isolated from trachoma patients in Sudan in 1963 was antigenically identical to an isolate from Saudi Arabia (A/SA1). No contemporary ocular C. trachomatis whole genome sequences have been reported from Sudan. METHODS: This study sequenced twenty ocular C. trachomatis isolates to improve understanding of pathogen diversity in North-East Africa and examine for genomic variation specific to Sudan, possibly related to the persistence of trachoma in surveyed communities. High quality, whole genome sequences were obtained from 12/20 isolates. RESULTS: All isolates were serovar A and had tarP and trpA sequences typical of classical, ocular C. trachomatis isolates. The Sudanese isolates formed a closely related subclade within the T2-trachoma clade of C. trachomatis phylogeny distinct from geographically disparate ocular isolates, with little intra-population diversity. We found 333 SNPs that were conserved in Sudanese ocular isolates but rare compared to other ocular C. trachomatis populations, which were focused in two genomic loci (CTA0172-CTA0173 and CTA0482). CONCLUSIONS: Limited intra-population diversity and geographical clustering of ocular C. trachomatis suggests minimal transmission between and slow diversification within trachoma-endemic communities. However, diversity may have been higher pre-treatment in these communities. Over-representation of Sudan-specific SNPs in three genes suggests they may have an impact on C. trachomatis growth and transmission in this population.

CCL17/TARC and CCR4 expression in Merkel cell carcinoma.

Merkel cell carcinoma (MCC) is a rare, highly aggressive neuroendocrine skin cancer. In more than 80% of the cases, Merkel cell polyomavirus (MCPyV) is a causal factor. The oncogenic potential of MCPyV is mediated through its viral oncoproteins, large T antigen (LT) and small t antigen (sT). To investigate the role of cytokines in MCC, a PCR array analysis for genes encoding inflammatory cytokines and receptors was performed on MCPyV-negative and MCPyV-positive MCC cell lines, respectively. We detected an increased expression of CCL17/TARC in the MCPyV-positive MKL2 cell line compared to the MCPyV-negative MCC13 cell line. Transfection studies in MCC13 cells with LT expression plasmid, and a luciferase reporter plasmid containing the CCL17/TARC promoter, exhibited stimulated promoter activity. Interestingly, the ectopic expression of CCL17/TARC upregulated MCPyV early and late promoter activities in MCC13 cells. Furthermore, recombinant CCL17/TARC activated both the mitogen-activated protein kinase and the NF-κB pathways. Finally, immunohistochemical staining on human MCC tissues showed a strong staining of CCL17/TARC and its receptor CCR4 in both LT-positive and -negative MCC. Taken together, CCL17/TARC and CCR4 may be a potential target in MCC therapy providing MCC patients with a better overall survival outcome.

The role of Merkel cell polyomavirus and other human polyomaviruses in emerging hallmarks of cancer.

Polyomaviruses are non-enveloped, dsDNA viruses that are common in mammals, including humans. All polyomaviruses encode the large T-antigen and small t-antigen proteins that share conserved functional domains, comprising binding motifs for the tumor suppressors pRb and p53, and for protein phosphatase 2A, respectively. At present, 13 different human polyomaviruses are known, and for some of them their large T-antigen and small t-antigen have been shown to possess oncogenic properties in cell culture and animal models, while similar functions are assumed for the large T- and small t-antigen of other human polyomaviruses. However, so far the Merkel cell polyomavirus seems to be the only human polyomavirus associated with cancer. The large T- and small t-antigen exert their tumorigenic effects through classical hallmarks of cancer: inhibiting tumor suppressors, activating tumor promoters, preventing apoptosis, inducing angiogenesis and stimulating metastasis. This review elaborates on the putative roles of human polyomaviruses in some of the emerging hallmarks of cancer. The reciprocal interactions between human polyomaviruses and the immune system response are discussed, a plausible role of polyomavirus-encoded and polyomavirus-induced microRNA in cancer is described, and the effect of polyomaviruses on energy homeostasis and exosomes is explored. Therapeutic strategies against these emerging hallmarks of cancer are also suggested.

Mixed venous versus central venous oxygen saturation in patients undergoing on pump beating coronary artery bypass grafting.

OBJECTIVE: To examine the validity of central venous oxygen saturation (ScvO(2)) as a numerical substitution of mixed venous oxygen saturation (SvO(2)) in adult patients undergoing normothermic on pump beating coronary artery bypass grafting (CABG). MATERIALS AND METHODS: Prospective clinical observational study was done at King Khalid University Hospital, King Saud University, Riyadh, Kingdom of Saudi Arabia. Thirty four adult patients scheduled for coronary artery surgery were included. Patients were monitored by a pulmonary artery catheter (PAC) as a part of our routine intraoperative monitoring. SvO(2) and ScvO(2) were simultaneously measured 15 minutes (T1) and 30 minutes (T2) after induction of anesthesia, 15 and 30 minutes after initiation of cardiopulmonary bypass (T3 and T4), and 15 and 30 minutes after admission to intensive care unit (T5 and T6). RESULTS: ScvO(2) showed higher reading than SvO(2) all through our study. Our results showed perfect positive statistically significant correlation between SvO(2) and ScvO(2) at all data points. Individual mean of difference (MOD) between both the readings at study time showed MOD of 1.34 and 1.44 at T1 and T2 simultaneously. This MOD was statistically insignificant, but after on pump beating normothermic bypass was initiated; MOD was 5.2 and 4.4 at T3 and T4 with high statistical significance. In ICU, MOD continues to have high statistical significance, MOD was 6.3 at T5 and at T6 it was 4.6. CONCLUSIONS: In on pump beating CABG patients; ScvO(2) and SvO(2) are not interchangeable numerically. ScvO(2) is useful in the meaning of trend; our data suggest that ScvO(2) is equivalent to SvO(2) , only in the course of clinical decisions as long as absolute values are not required.