Generation of an induced pluripotent stem cell line from a patient with epileptic encephalopathy caused by the CYFIP2 R87C variant.

Induced pluripotent stem cells (iPSCs) opened the possibility to use patient cells as a model for several diseases. iPSCs can be reprogrammed from somatic cells collected in a non-invasive way, and then differentiated into any other cell type, while maintaining the donor´s genetic background. CYFIP2 variants were associated with the onset of an early form of epileptic encephalopathy. Studies with patients showed that the R87C variant seems to be one of the variants that causes more severe disease, however, to date there are no studies with a human cell model that allows investigation of the neuronal phenotype of the R87C variant. Here, we generated an iPSC line from a patient with epileptic encephalopathy caused by the CYFIP2 R87C variant. We obtained iPSC clones by reprogramming urinary progenitor cells from a female patient. The generated iPSC line presented a pluripotent stem cell morphology, normal karyotype, expressed pluripotency markers and could be differentiated into the three germ layers. In further studies, this cell line could be used as model for epileptic encephalopathy disease and drug screening studies.

Accessing the Human Pluripotent Stem Cell Translatome by Polysome Profiling.

Polysome profiling is a technique that uses sucrose density gradient ultracentrifugation to separate complexes of mRNAs associated with one or more ribosomes. Here we describe polysome profiling analysis in human pluripotent stem cells (hPSCs) using a continuous ultraviolet spectrophotometer and a gradient fractionator. We provide protocols for processing sucrose gradient fractions for isolation of RNA for RT-qPCR or large-scale sequencing analysis, used to establish the translational status of specific mRNAs and identify the role of noncoding RNA in translation.

Investigation of anti-inflammatory and anti-proliferative activities promoted by photoactivated cationic porphyrin.

BACKGROUND: Photodynamic therapy (PDT) is a technique that uses light and a photosensitizer, converting local molecular oxygen into singlet oxygen, which eliminates a target unhealthy tissue. It has been increasingly used for the treatment of several diseases including skin disorders. Psoriasis is a chronic inflammatory skin disease expressing immune and hyperproliferative features. OBJECTIVE: This study aimed to evaluate the effect of the photosensitizer 5,10-diphenyl-15,20-di(N-methylpyridinium-4-yl)porphyrin (Di-cis-Py+) in in vivo models whereby some psoriasis-like parameters could be investigated. METHODS: The antiinflammation and antiproliferative activities of Di-cis-Py+ photoactivated was measured by myeloperoxidase (MPO) and N-acetyl-ß-d-glucosaminidase (NAG) enzyme activity assay, measurement of IL-6, IL-1ß and TNF-α levels, evaluation of proliferating cell nuclear antigen (PCNA) levels by immunohistochemistry and by Western blot. RESULTS: Treatment involving PDT and Di-cis-Py+ resulted in reduction of edema, cellular infiltration, proinflammatory cytokines, as well as reduced hyperproliferation of the epidermis. All the evaluated parameters were promoted by topical application of phlogistic agents and are similar to that observed in lesions of psoriatic skin. CONCLUSION: The results shows the advantage of topical application, do not cause apparently photosensitivity and have effects comparable to dexamethasone, a first-line drug for the treatment of the disease.