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Background: Pharmacokinetic study plays important role in evaluating of the druggability of investigational drugs and clinical monitoring of marketed drugs. Methods: This work developed a sensitive chemiluminescence (CL) method for rapidly studying drug pharmacokinetics in rats using matrine as a probe. The method involved a flow-injection CL (FI-CL) technique that enables matrine to inhibit a luminol/sulfobutylether-β-cyclodextrin (SBE-β-CD)- CL reaction system. Results: A good linear relationship was found between the inhibition of CL intensity and the logarithm of matrine concentration over the range of 0.28 ng/ml to 560.0 ng/ml with a detection limit of 0.1 ng/ml (3ó). The maximum drug plasma concentration (Cmax), time to Cmax, absorption half-life (t1/2α), elimination half-life (t1/2β), and areas under the plasma concentration-time curve AUC(0-t) and AUC(0-∞) were determined to be 48.29±2.45 µg/ml, 0.083±0.006 h, 0.079±0.005 h, 0.079±0.03 h, 23.79±1.16 µg/h•ml and 30.51±0.82 µg/h•ml when the rats were administrated matrine by intravenous injection through tail. The values of(t1/2α), t1/2β, AUC(0-t) and AUC(0-∞) changed to 0.073±0.005 h, 0.82±0.03 h, 32.03±1.31 µg/h•ml and 35.24±0.92 µg/h•ml when the dosed matrine was replaced by SBE-β- CD/matrine inclusion. The pharmacokinetic profile of matrine was in accordance with an open twocompartment model after administration of matrine alone or SBE-β-CD/matrine inclusion. These results indicated that SBE-β-CD inclusion sustained the release of matrine, prolonged the matrine distribution from the blood to the tissues and improved its bioavailability. Conclusions: The proposed method has potential to become a powerful alternative for rapid pharmacokinetic studies due to the advantages of a good linear range, high sensitivity, high recovery and superior analytical efficiency.

Methods for the in vitroculture of primary small intestinal epithelium have improved greatly in recent years. A critical barrier for the translation of this methodology to the patient's bedside is the ability to grow intestinal stem cells using a well-defined extracellular matrix. Current methods rely on the use of Matrigel™, a proprietary basement membrane-enriched extracellular matrix gel produced in mice that is not approved for clinical use. We demonstrate for the first time the capacity to support the long-term in vitrogrowth of murine intestinal epithelium in monoculture, using type I collagen. We further demonstrate successful in vivoengraftment of enteroids co-cultured with intestinal subepithelial myofibroblasts in collagen gel. Small intestinal crypts were isolated from 6 to 10 week old transgenic enhanced green fluorescent protein (eGFP+) mice and suspended within either Matrigel or collagen gel; cultures were supported using previously reported media and growth factors. After 1 week, cultures were either lysed for DNA or RNA extraction or were implanted subcutaneously in syngeneic host mice. Quantitative real-time polymerase chain reaction (qPCR) was performed to determine expansion of the transgenic eGFP-DNA and to determine the mRNA gene expression profile. Immunohistochemistry was performed on in vitrocultures and recovered in vivoexplants. Small intestinal crypts reliably expanded to form enteroids in either Matrigel or collagen in both mono- and co-cultures as confirmed by microscopy and eGFP-DNA qPCR quantification. Collagen-based cultures yielded a distinct morphology with smooth enteroids and epithelial monolayer growth at the gel surface; both enteroid and monolayer cells demonstrated reactivity to Cdx2, E-cadherin, CD10, Periodic Acid-Schiff, and lysozyme. Collagen-based enteroids were successfully subcultured in vitro, whereas pure monolayer epithelial sheets did not survive passaging. Reverse transcriptase-polymerase chain reaction demonstrated evidence of Cdx2, villin 1, mucin 2, chromogranin A, lysozyme 1, and Lgr5 expression, suggesting a fully elaborated intestinal epithelium. Additionally, collagen-based enteroids co-cultured with myofibroblasts were successfully recovered after 5 weeks of in vivoimplantation, with a preserved immunophenotype. These results indicate that collagen-based techniques have the capacity to eliminate the need for Matrigel in intestinal stem cell culture. This is a critical step towards producing neo-mucosa using good manufacturing practices for clinical applications in the future.

Poly (TBAM-AM-NMMAD) random-block copolymer were prepared by aqueous micellar copolymerization using AIBA (V50), ammonium persulfate (J) and sodium bisulfite (D) as initiator and SDS as surfactant in the mixture solvent of deionized water. The structure of Poly (TBAM-AM-NMMAD) was characterized by solid state 13C-NMR and FTIR spectroscopy and the dynamic viscosity was measured by the NDJ-1 viscometer. The experimental results and analysis shows poly (AM-TBAM-NMMAD) random-block copolymer which had been endowed with high rheological properties was successfully synthesized by controlling the abundance of initiator based on the method of orthogonal experiment.

Human small intestinal crypts are the source of intestinal stem cells (ISCs) that are capable of undergoing self-renewal and differentiation to an epithelial layer. The development of methods to expand the ISCs has provided opportunities to model human intestinal epithelial disorders. Human crypt samples are usually obtained from either endoscopic or discarded surgical samples, and are thereby exposed to warm ischemia, which may impair their in vitrogrowth as three-dimensional culture as spheroids or enteroids. In this study we compared duodenal samples obtained from discarded surgical samples to those isolated from whole-body preserved cadaveric donors to generate in vitrocultures. We also examined the effect of storage solution (phosphate-buffered saline or University of Wisconsin [UW] solution) as well as multiple storage times on crypt isolation and growth in culture. We found that intestinal crypts were successfully isolated from cadaveric tissue stored for up to 144 h post-procurement and also were able to generate enteroids and spheroids in certain media conditions. Surgical samples stored in UW after procurement were sufficiently viable up to 24 h and also allowed the generation of enteroids and spheroids. We conclude that surgical samples stored for up to 24 h post-procurement in UW solution allowed for delayed crypt isolation and viable in vitrocultures. Furthermore, in situ, hypothermic preservation in cadaveric duodenal samples permitted crypt/ISC isolation, and successful culture of spheroids and enteroids from tissues held for up to 6 days post-procurement.