A surface-tethered spheroid model for functional evaluation of 3T3-L1 adipocytes.

In order to effectively treat obesity, it must be better understood at the cellular level with respect to metabolic state and environmental stress. However, current two-dimensional (2D) in vitro cell culture methods do not represent the in vivo adipose tissue appropriately due to the absence of complex architecture and cellular signaling. Conversely, 3D in vitro cultures have been reported to have optimal results mimicking the adipose tissue in vivo. The main aim of this study was to examine the efficacy of a novel conjugate of a genetically engineered polymer, elastin-like polypeptide (ELP) and a synthetic polymer, polyethyleneimine (PEI), toward creating a 3D preadipocyte culture system. We then used this 3D culture model to study the preadipocyte differentiation and adipocyte maintenance processes when subjected to various dosages of nutritionally relevant free fatty acids with respect to total DNA and protein content, cell viability, and intracellular triglyceride accumulation. Our results showed that 3T3-L1 preadipocytes cultured on the ELP-PEI surface formed 3D spheroids within 72 h, whereas the cells cultured on unmodified tissue culture polystyrene surfaces remained in monolayer configuration. Significant statistical differences were discovered between the 3D spheroid and 2D monolayer culture with respect to the DNA and protein content, fatty acid consumption, and triglyceride accumulation, indicating differences in cellular response. Results indicated that the 3D culture may be a more sensitive modeling technique for in vitro adipocyte culture and provides a platform for future evaluation of 3D in vitro adipocyte function.

Effect of processing temperature on the morphology and drug-release characteristics of elastin-like polypeptide-collagen composite coatings.

Elastin-like polypeptides (ELPs) exhibit an inverse phase transition temperature (Tt) in response to changes in their environment. We hypothesized that processing ELP-collagen composites at temperatures higher than the Tt of ELP (∼32 °C) will affect their microstructure and subsequently, achieve tunable release of model drugs. The composite coatings were prepared by formation of ELP-collagen hydrogels at 37 °C, incubation at 37, 45, or 55 °C, and finally air-drying at 37 °C. Scanning electron micrographs revealed that the fabrication process affected both the collagen and ELP microaggregate phases. A gradual time dependent bovine serum albumin (BSA) release that followed the power law and a burst antibiotic doxycycline release followed by a linear zero-order release were observed. Importantly, BSA and doxycycline releases were dependent on the ELP microaggregate size, which was governed by the processing temperatures. This study lays the foundation to achieve optimized composite microstructures by controlling processing conditions for drug delivery applications.

Vitamin D Supplementation Suppresses Hypoxia-Stimulated Placental Cytokine Secretion, Hypertension and CD4+ T Cell Stimulation in Response to Placental Ischemia.

OBJECTIVE: To investigate a role of Vitamin D in the pathogenesis of preeclampsia (PE), and to discern any potential benefits of Vitamin D supplementation on hypertension in the RUPP rat model of PE. STUDY DESIGN: Blood and placentas from normal pregnancies (NP) and PE were collected following elective cesarean delivery without evidence of infection. Circulating Vitamin D was extracted by HPLC and measured via mass spectrometry. Media for placenta explants was supplemented with Vitamin D and exposed to hypoxic (1% O2) or normoxic (6% O2) conditions for 24 hours. ELISAs were performed on media and normalized to total protein to determine cytokine secretion. RUPP rats were supplemented with vitamin D by oral gavage, and blood pressure (MAP) and pup weights were measured in NP and RUPP rats with or without Vitamin D supplementation. Flow cytometry was used to evaluate CD4+ Tcells in control RUPP rats and RUPP rats treated with Vitamin D. RESULTS: Inflammatory cytokine secretion was higher (p<0.05) while the anti-inflammatory cytokine, IL-10, was significantly lower in the media of PE placentas compared to NP (p=0.005). Vitamin D supplementation decreased hypoxia stimulated pro-inflammatory cytokine secretion (p=0.003) in the media of PE placentas. Vitamin D decreased MAP and circulating CD4+ T cells in the RUPP rat model of PE (p<0.05). CONCLUSION: Vitamin D supplementation may be useful in the treatment or prevention of hypertensive disorders in pregnancy.