Screening fibroblasts

In vitro studies on isolated cells including hepatocytes, erythrocytes, fibroblasts, and alveolar cells continue to demonstrate the specificity of action that these toxins have for liver cells (83,86,93). This specificity has led Aune and Berg (94) to use isolated rat hepatocytes as a screen for detecting hepatotoxic waterblooms of cyanobacteria. 

In many localities, newborn screening has become standard for this disorder, which in the general population has an approximate incidence of 1/250,000 live births. Carrier detection is possible, either by measurement of enzymatic activity in cultured fibroblasts or by study of restriction endonuclease fragments of DNA. Antenatal testing is also available. 

A much more serious genetic disease, first described by Foiling in 1934, is phenylketonuria. Here the disturbance in phenylalanine metabolism is due to an autosomal recessive deficiency in liver phenylalanine hydroxylase (Jervis, 1954) which normally converts significant amounts of phenylalanine to tyrosine. Phenylalanine can therefore only be metabolized to phenylpyruvate and other derivatives, a route which is inadequate to dispose of all the phenylalanine in the diet. The amino acid and phenylpyruvate therefore accummulate. The condition is characterized by serious mental retardation, for reasons which are unknown. By the early 1950s it was found that if the condition is diagnosed at birth and amounts of phenylalanine in the diet immediately and permamently reduced, mental retardation can be minimized. The defect is shown only in liver and is not detectable in amniotic fluid cells nor in fibroblasts. A very sensitive bacterial assay has therefore been developed for routine screening of phenylalanine levels in body fluids in newborn babies. 

Polarographic studies can also be used to screen for much more specific defects suspected in some patients, such as impaired mitochondrial glutamate transport in cultured skin fibroblasts from patients presenting with neonatal myoclonic epilepsy [64]. 

All of the above-mentioned patterns are specific for the particular disease. The method is suitable as an initial screen to identify those patients in whom such a disorder must be excluded. The diagnosis must then be confirmed by enzyme analysis in serum, leucocytes or cultured skin fibroblasts or by way of mutation analysis. 

The test is performed for diagnosis of all clinical forms of SASD. This analysis is usually done after an initial TLC screening test that is positive for free sialic acid, and an increased free sialic acid value in the quantitative urine determination test. The test is like the quantitative urine test performed with the periodate-TBA assay [5, 22]. However, in this case interference is decreased by prepurification of the sample using ion-exchange chromatography [12]. Fibroblasts are cultured under standardized conditions. Cell lysates are prepared by tip sonification in distilled water and the cleared lysates are applied to small Dowex columns. NeuAc is eluted, freeze dried,... 

Cell-based systems are biology s way of dividing the expressed genome into functional units. In drag discovery, a focus on specific cell systems, such as T-cells and fibroblasts for assembly of human immunodeficiency vims or herpes simplex vims particles and endothelial cells for demonstrating adhesion-dependent processes, often provides convenient primary dmg screening systems. 

A sublibrary L17, made of 42 diverse library individuals (Fig. 11.23), was screened in an in vivo model for the proliferation of fibroblasts, where a high value represented a candidate for the tissue reconstruction process around the medical implant. The screening protocol was performed in parallel, and a correlation was established between the fibroblast proliferation (biological property) and the air-water contact angle (physicochemical property). A crude SAR was assessed for further, more... 

Indoprofen In a high-throughput screen of ca. 47,000 compound library, authors [43] have converged on indoprofen as a molecule that enhanced production of an SMN2- vs. SMN1 -luciferase reporter protein. Indoprofen, a NSAID and cyclooxygenase (COX) inhibitor, afforded a 13% enhancement of Smn protein and a fivefold increase in the number of nuclear gems in fibroblasts from SMA patients. Notably, other tested NSAIDs or COX inhibitors were inactive in the assay. 

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