COS-7-based model: methodological approach to study John Cunningham virus replication cycle.

John Cunningham virus (JCV) is a human neurotropic polyomavirus whose replication in the Central Nervous System (SNC) induces the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). JCV propagation and PML investigation have been severely hampered by the lack of an animal model and cell culture systems to propagate JCV have been very limited in their availability and robustness. We previously confirmed that JCV CY strain efficiently replicated in COS-7 cells as demonstrated by the progressive increase of viral load by quantitative PCR (Q-PCR) during the time of transfection and that archetypal regulatory structure was maintained, although two characteristic point mutations were detected during the viral cycle. This short report is an important extension of our previous efforts in defining our reliable model culture system able to support a productive JCV infection.Supernatants collected from transfected cells have been used to infect freshly seeded COS-7 cell line. An infectious viral progeny was obtained as confirmed by Western blot and immunofluorescence assay. During infection, the archetype regulatory region was conserved.Importantly, in this study we developed an improved culture system to obtain a large scale production of JC virus in order to study the genetic features, the biology and the pathogenic mechanisms of JC virus that induce PML.

Dynamic regulation of the cancer stem cell compartment by Cripto-1 in colorectal cancer.

Stemness was recently depicted as a dynamic condition in normal and tumor cells. We found that the embryonic protein Cripto-1 (CR1) was expressed by normal stem cells at the bottom of colonic crypts and by cancer stem cells (CSCs) in colorectal tumor tissues. CR1-positive populations isolated from patient-derived tumor spheroids exhibited increased clonogenic capacity and expression of stem-cell-related genes. CR1 expression in tumor spheroids was variable over time, being subject to a complex regulation of the intracellular, surface and secreted protein, which was related to changes of the clonogenic capacity at the population level. CR1 silencing induced CSC growth arrest in vitro with a concomitant decrease of Src/Akt signaling, while in vivo it inhibited the growth of CSC-derived tumor xenografts and reduced CSC numbers. Importantly, CR1 silencing in established xenografts through an inducible expression system decreased CSC growth in both primary and metastatic tumors, indicating an essential role of CR1 in the regulation the CSC compartment. These results point to CR1 as a novel and dynamically regulated effector of stem cell functions in colorectal cancer.

Patient comorbidity: relationship to outcomes of total knee arthroplasty.

One hundred six patients treated consecutively with total knee arthroplasty were evaluated to determine whether preoperative comorbidity (as measured by patient class, knee score, short form, anesthesia severity assessment, and number of medical comorbidities) correlated with perioperative and postoperative outcomes, including length of stay, total (and specific) hospital charges, and validated outcome scores. The length of stay for total knee arthroplasty was longer in patients who had lower preoperative knee scores and for patients with greater medical and musculoskeletal morbidity. Greater total hospital costs were associated with Class C patients and patients with poor anesthesia morbidity ratings. Patients who were debilitated medically and had four or more risk factors had decreased postoperative outcome scores. Preoperative medical and musculoskeletal morbidity influence the results of total knee arthroplasty. These findings may be useful to surgeons for optimizing resource utilization and outcomes in patients undergoing total knee arthroplasty. These data must be accounted for when contrasting total knee arthroplasty results between different surgeons and institutions.