Steroidogenic enzymes in leydig cells

A number of investigators have reported on a rather rare syndrome of excess aromatase activity. In boys, it can lead to gynecomastia , and in girls to precocious puberty and gigantomastia . In both sexes, early epiphyseal closure leads to short stature. This condition is due to mutations in the CYP19A1 gene which encodes aromatase. [16] It is inherited in an autosomal dominant fashion. [17] It has been suggested that the pharaoh Akhenaten and other members of his family may have suffered from this disorder, [18] but more recent genetic tests suggest otherwise. [19] It is one of the causes of familial precocious puberty—a condition first described in 1937. [20]

The StAR protein was first identified, characterized and named by Dr. Douglas Stocco at Texas Tech University Health Sciences Center in 1994. [18] The role of this protein in lipoid CAH was confirmed the following year in collaboration with Dr. Walter Miller at the University of California, San Francisco . [19] All of this work follows the initial observations of the appearance of this protein and its phosphorylated form coincident with factors that caused steroid production by Dr. Nanette Orme-Johnson while at Tufts University . [20]

Steroid isolation , depending on context, is the isolation of chemical matter required for chemical structure elucidation, derivitzation or degradation chemistry, biological testing, and other research needs (generally milligrams to grams, but often more [37] or the isolation of "analytical quantities" of the substance of interest (where the focus is on identifying and quantifying the substance (for example, in biological tissue or fluid). The amount isolated depends on the analytical method, but is generally less than one microgram. [38] [ page needed ] The methods of isolation to achieve the two scales of product are distinct, but include extraction , precipitation, adsorption , chromatography , and crystallization . In both cases, the isolated substance is purified to chemical homogeneity; combined separation and analytical methods, such as LC-MS , are chosen to be "orthogonal"—achieving their separations based on distinct modes of interaction between substance and isolating matrix—to detect a single species in the pure sample. Structure determination refers to the methods to determine the chemical structure of an isolated pure steroid, using an evolving array of chemical and physical methods which have included NMR and small-molecule crystallography . [2] :10–19 Methods of analysis overlap both of the above areas, emphasizing analytical methods to determining if a steroid is present in a mixture and determining its quantity. [38]

Leydig cell isolation and culture. Leydig cells were isolated as described previously ( Klinefelter et al. 1987 ), with slight modifications. The testes were decapsulated, placed in a dissociation buffer [M-199 medium with g/L HEPES, g/L bovine serum albumin (BSA), g/L sodium bicarbonate (pH ), and 25 mg/L trypsin inhibitor] containing collagenase ( mg/mL) at 34°C, and shaken for 30 min. Digested testes were passed through a 100-μm nylon mesh, and Leydig cells were purified by Percoll gradient separation. The final purity of the Leydig cells, determined by staining the cells for 3β-HSD activity, was consistently approximately 90%.

Steroidogenic enzymes in leydig cells

steroidogenic enzymes in leydig cells

Leydig cell isolation and culture. Leydig cells were isolated as described previously ( Klinefelter et al. 1987 ), with slight modifications. The testes were decapsulated, placed in a dissociation buffer [M-199 medium with g/L HEPES, g/L bovine serum albumin (BSA), g/L sodium bicarbonate (pH ), and 25 mg/L trypsin inhibitor] containing collagenase ( mg/mL) at 34°C, and shaken for 30 min. Digested testes were passed through a 100-μm nylon mesh, and Leydig cells were purified by Percoll gradient separation. The final purity of the Leydig cells, determined by staining the cells for 3β-HSD activity, was consistently approximately 90%.

Media:

steroidogenic enzymes in leydig cellssteroidogenic enzymes in leydig cellssteroidogenic enzymes in leydig cellssteroidogenic enzymes in leydig cellssteroidogenic enzymes in leydig cells