Chromatin status and transcription factor binding to gonadotropin promoters in gonadotrope cell lines.

BACKGROUND: Proper expression of key reproductive hormones from gonadotrope cells of the pituitary is required for pubertal onset and reproduction. To further our understanding of the molecular events taking place during embryonic development, leading to expression of the glycoproteins luteinizing hormone (LH) and follicle-stimulating hormone (FSH), we characterized chromatin structure changes, imparted mainly by histone modifications, in model gonadotrope cell lines. METHODS: We evaluated chromatin status and gene expression profiles by chromatin immunoprecipitation assays, DNase sensitivity assay, and RNA sequencing in three developmentally staged gonadotrope cell lines, αT1-1 (progenitor, expressing Cga), αT3-1 (immature, expressing Cga and Gnrhr), and LßT2 (mature, expressing Cga, Gnrhr, Lhb, and Fshb), to assess changes in chromatin status and transcription factor access of gonadotrope-specific genes. RESULTS: We found the common mRNA α-subunit of LH and FSH, called Cga, to have an open chromatin conformation in all three cell lines. In contrast, chromatin status of Gnrhr is open only in αT3-1 and LßT2 cells. Lhb begins to open in LßT2 cells and was further opened by activin treatment. Histone H3 modifications associated with active chromatin were high on Gnrhr in αT3-1 and LßT2, and Lhb in LßT2 cells, while H3 modifications associated with repressed chromatin were low on Gnrhr, Lhb, and Fshb in LßT2 cells. Finally, chromatin status correlates with the progressive access of LHX3 to Cga and Gnrhr, followed by PITX1 binding to the Lhb promoter. CONCLUSION: Our data show the gonadotrope-specific genes Cga, Gnrhr, Lhb, and Fshb are not only controlled by developmental transcription factors, but also by epigenetic mechanisms that include the modulation of chromatin structure, and histone modifications.

Effects of Dissolution Medium pH and Simulated Gastrointestinal Contraction on Drug Release From Nifedipine Extended-Release Tablets.

In contrast to nifedipine matrix-based extended-release dosage forms, the osmotic pump drug delivery systems have a zero-order drug release independent of external variables such as pH, agitation rate, and dissolution media. The objective of this study focuses on the in vitro evaluation of the mechanical properties of osmotic pump and polymer matrix-based formulations in dissolution media, and the potential impacts that media pH and simulated gastrointestinal contraction have on drug release. Two strengths of osmotic pump product A and polymer matrix-based product B were used in this study. An in-house system was developed with the capability of applying mechanical compression and monitoring mechanical properties of sample during dissolution testing. A United States Pharmacopeia or an in-house apparatus was used for dissolution testing under various conditions. Compared to the product A, the mechanical properties of the product B change significantly at various pHs and mechanical compressions. The results suggest that polymer matrix-based products bear a risk of formulation-related interactions with the gastrointestinal tract during in vivo drug dissolution, especially in the case of concomitant pH and gastric contractile changes. Modified dissolution testing devices may help formulation scientists in product development and provide regulatory agencies with an additional metric for quality assurance of drug products.