Control of self-renewal and differentiation of human ES cells (hESCs)remains a challenge. This is largely due to the use of culture systems that involve poorly defined animal products and do not mimic the normal developmental milieu. Routine protocols involve the propagation of hESCs on mouse fibroblast or human feeder layers, enzymatic cell removal, and spontaneous differentiation in cultures of embryoid bodies, and each of these steps involves significant variability of culture conditions. We report that a completely synthetic hydrogel matrix can support (i) long-term self-renewal of hESCs in the presence of conditioned medium from mouse embryonic fibroblast feeder layers, and (ii) direct cell differentiation. Hyaluronic acid (HA) hydrogels were selected because of the role of HA in early development and feeder layer cultures of hESCs and the controllability of hydrogel architecture, mechanics, and degradation. When encapsulated in 3D HA hydrogels (but not within other hydrogels or in monolayer cultures on HA), hESCs maintained their undifferentiated state, preserved their normal karyotype, and maintained their full differentiation capacity as indicated by embryoid body formation. Differentiation could be induced within the same hydrogel by simply altering soluble factors. We therefore propose that HA hydrogels, with their developmentally relevant composition and tunable physical properties, provide a unique microenvironment for the selfrenewal and differentiation of hESCs.scaffolds ͉ three-dimensional cultures ͉ vasculogenesis

In vitro models of the liver using isolated primary hepatocytes have been used as screens for measuring the metabolism, toxicity and efficacy of xenobiotics, for studying hepatocyte proliferation, and as bioartificial liver support systems. Yet, primary isolated hepatocytes rapidly lose liver specific functions when maintained under standard in vitro cell culture conditions. Many modifications to conventional culture methods have been developed to foster retention of hepatocyte function. Still, not all of the important functions -- especially the biotransformation functions of the liver -- can as yet be replicated at desired levels, prompting continued development of new culture systems. In the first part of this article, we review primary hepatocyte in vitro systems used in metabolism and enzyme induction studies. We then describe a scalable microreactor system that fosters development of 3D-perfused micro-tissue units and show that primary rat cells cultured in this system are substantially closer to native liver compared to cells cultured by other in vitro methods, as assessed by a broad spectrum of gene expression, protein expression and biochemical activity metrics. These results provide a foundation for extension of this culture model to other applications in drug discovery -- as a model to study drug-drug interactions, as a model for the assessment of acute and chronic liver toxicity arising from exposure to drugs or environmental agents; and as a disease model for the study of viral hepatitis infection and cancer metastasis.

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The use of nanoparticles for targeted drug delivery is often facilitated by specific conjugation of functional targeting molecules to the nanoparticle surface. We compared different biotin binding proteins (avidin, streptavidin, or neutravidin) as crosslinkers to conjugate proteins to biodegradable nanoparticles prepared from PLGA-PEG-biotin polymers. Avidin gave the highest levels of overall protein conjugation, whereas neutravidin minimized protein non-specific binding to the polymer. The tetanus toxin C fragment (TTC), which is efficiently retrogradely transported in neurons and binds to neurons with high specificity and affinity, retained the ability to bind to neuroblastoma cells following amine group modifications. TTC was conjugated to nanoparticles using neutravidin, and the resulting nanoparticles were shown to selectively target neuroblastoma cells in vitro. TTCconjugated nanoparticles have the potential to serve as drug delivery vehicles targeted to the central nervous system.

Abstract-TheUse of specific adrenergic receptor knockout (KO) mice has also been useful in defining the role of ␣ 1 AR subtypes in cardiovascular regulation. [3][4][5][6] However, results in KO mice remain controversial, with each subtype contributing to regulation of blood pressure and vascular contractility but without a subtype-selective role being clearly apparent. The ␣ 1 A, B, and D AR subtypes have been shown to contribute to the maintenance of basal blood pressure and vasopressor responses to exogenous agonists. Specifically, ␣ 1A and ␣ 1D AR KO mice have lower basal blood pressure, whereas all 3 KO mice models have attenuated pressor responses to exogenous norepinephrine (NE). However, it is becoming increasingly appreciated that differential ␣ 1 AR-mediated responses may be the result of different receptor subtypes at junctional versus extrajunctional sites. 7-9 Thus, a critical and clinically relevant question in ␣ 1 AR adrenergic biology remains: is there a specific receptor subtype that mediates sympathetic transmission at the neuroeffector junction? 10 Clinical data suggest that orthostatic intolerance (OI) may result when ␣ 1B ARs are specifically inhibited. 11 Thus, we hypothesized that the ␣ 1B AR is the subtype critical in mediating vasoconstriction at the neuroeffector junction. To test the hypothesis, we measured integrated cardiovascular responses to selective carotid arterial baroreceptor unloading induced by transient bilateral carotid occlusion (TBCO) in mice with a homozygous deletion of the ␣ 1B AR gene. We also determined the vascular contractile responses in mesenteric resistance vessels in vitro to endogenous NE (mediated by electrical field stimulation [EFS]) in KO and wild-type (WT) mice.