Hepatocyte spheroid culture on a polydimethylsiloxane chip having microcavities

A two-dimensional microarray technique of spherical multicellular aggregates (spheroids) using a microfabricated polydimethylsiloxane (PDMS) chip and the expression of liver-specific functions of primary rat hepatocytes on the chip were investigated. The PDMS chip, which was fabricated by a photolithography-based technique, consisted of approximately 2500 cylindrical microcavities (approximately 1100 cavities/cm²) in a triangular arrangement of 330 μm pitch on a PDMS plate (20 × 20 mm); each cavity measured 300 μm in diameter and 100 μm in depth. Most hepatocytes on the PDMS chip gradually gathered and subsequently formed a single spheroid in each cavity until 3 days of culture. A part of the spheroid was attached to the bottom or wall surface of the microcavity, and the spheroid configuration was maintained for at least 14 days of culture. Albumin secretion, ammonia removal and ethoxyresorufin O-dealkylase (EROD) activity, which is a cytochrome P-450-dependent reaction, of hepatocytes on the PDMS chip were higher than those of a monolayer dish or a flat PDMS dish without microcavities, and were maintained for at least 10 days of culture. The spheroid microarray technique appears to be promising in the development of cell chips and microbioreactors.     

Oral health provisions in U.S. health care reform

BackgroundDuring 2009, both chambers of the U.S. Congress passed health care reform bills that contained a variety of provisions specific to oral health and dental care. In March 2010, the Senate version—the Patient Protection and Affordable Care Act (referred to as the Affordable Care Act [ACA])—was signed into law.MethodsThe authors establish the context for ACA dental provisions by reviewing prior federal legislation pertaining to dental coverage. They analyze the final U.S. House and Senate health care reform bills for their oral health content and draw observations regarding congressional interest in oral health.ResultsThe authors identify and describe more than 30 provisions of direct relevance to dentistry within the domains of insurance coverage, dental workforce, safety net, prevention and surveillance. Although the two bills differed in many details, both address oral health infrastructure and delivery of care, with particular attention to underserved child and adolescent populations.ConclusionsThe oral health provisions in the health care reform bills evidenced strong congressional interest in oral health and dental care, with an emphasis on equitable care for children.Practice ImplicationsThe effect of each congressional action on the future of dental practice will depend on how the provisions are regulated and implemented. The dental profession needs to recognize the strong and ongoing interest of lawmakers in oral health care and must maintain active engagement in the policymaking process.     

Carboxyl-terminus of Hsc70 interacting protein mediates 2,5-hexanedione-induced neurofilament medium chain degradation

Neurofilaments (NFs), the most abundant cytoskeletal components in large neurons and myelinated axons, are the targets of n-hexane-induced neuropathy, in which a specific loss of NFs protein has been frequently observed. However, the precise mechanisms regulating NFs contents are not well understood. The aim of this study was to elucidate the role of ubiquitin–proteasome system (UPS) in NFs degradation. We first demonstrated that the E3 ligase carboxyl-terminus of Hsc70 interacting protein (CHIP), originally identified as a co-chaperone of Hsc70, directly interacted with NFs medium chain (NF-M) and then enhanced NF-M ubiquitination and degradation after 2,5-hexanedione (HD) treatment. Consistent with this result, the application of proteasome inhibitor MG132 partly reversed HD-induced decrease of NF-M. Finally, we found that other components of UPS system (e.g. ubiquitin-activating enzyme E1, CHIP and proteasome) were significantly increased in sciatic nerve of HD-intoxicated rats. In conclusion, this study indicated that the CHIP ubiquitin ligase complex interacted with and repressed NFs by targeting NFs for ubiquitin-mediated proteolysis, which led to reduction of NFs contents in HD-induced neuropathy.     

组蛋白乙酰化酶调控MSCs经5-azaC诱导后的细胞周期和增殖特性

目的 阐述组蛋白乙酰化酶GCN5在间充质干细胞经5-氮杂胞苷诱导过程中的作用及其转录调控机制。方法 MTT法、流式细胞术检测细胞周期和细胞增殖;定量PCR检测P21基因表达;CHIP技术验证GCN5募集蛋白复合体与P21基因的相互作用及P21基因启动子区域的乙酰化水平;Co-IP技术分离、串联质谱技术鉴定GCN5募集蛋白复合体组成。结果 MSCs经5-azaC诱导3 d后G0/G1期细胞比例、P21基因表达量最高,此后逐渐降低,细胞增殖指数及G2/S期细胞比例与上述结果相反;筛选出GCN5募集蛋白复合体为：依赖ATP的染色质重塑复合体成分、转录起始复合体成分、转录因子和锌指结构蛋白;诱导组GCN5与P21基因启动子区域结合能力及P21基因启动子区域组蛋白H3乙酰化水平高于未诱导组。结论 GCN5募集蛋白复合体通过结合于P21基因启动子区域参与调控MSCs经5-azaC诱导体外分化过程中细胞周期G0/G1期和细胞增殖特性的调控。     

Heat shock proteins: Cellular and molecular mechanisms in the central nervous system

Emerging evidence indicates that heat shock proteins (HSPs) are critical regulators in normal neural physiological function as well as in cell stress responses. The functions of HSPs represent an enormous and diverse range of cellular activities, far beyond the originally identified roles in protein folding and chaperoning. HSPs are now understood to be involved in processes such as synaptic transmission, autophagy, ER stress response, protein kinase and cell death signaling. In addition, manipulation of HSPs has robust effects on the fate of cells in neurological injury and disease states. The ongoing exploration of multiple HSP superfamilies has underscored the pluripotent nature of HSPs in the cellular context, and has demanded the recent revamping of the nomenclature referring to these families to reflect a re-organization based on structure and function. In keeping with this re-organization, we first discuss the HSP superfamilies in terms of protein structure, regulation, expression and distribution in the brain. We then explore major cellular functions of HSPs that are relevant to neural physiological states, and from there we discuss known and proposed HSP impacts on major neurological disease states. This review article presents a three-part discussion on the array of HSP families relevant to neuronal tissue, their cellular functions, and the exploration of therapeutic targets of these proteins in the context of neurological diseases.     

Using Appendicitis to Improve Estimates of Childhood Medicaid Participation Rates

Objective

Administrative data are often used to estimate state Medicaid/Children's Health Insurance Program duration of enrollment and insurance continuity, but they are generally not used to estimate participation (the fraction of eligible children enrolled) because administrative data do not include reasons for disenrollment and cannot observe eligible never-enrolled children, causing estimates of eligible unenrolled to be inaccurate. Analysts are therefore forced to either utilize survey information that is not generally linkable to administrative claims or rely on duration and continuity measures derived from administrative data and forgo estimating claims-based participation. We introduce appendectomy-based participation (ABP) to estimate statewide participation rates using claims by taking advantage of a natural experiment around statewide appendicitis admissions to improve the accuracy of participation rate estimates.

The neuronal ubiquitin-proteasome system: murine models and their neurological phenotype

The ubiquitin-proteasome system (UPS) is the main intracellular pathway for regulated protein turnover. This system is of vital importance for maintaining cellular homeostasis and is essential for neuronal functioning. It is therefore not surprising that impairment of this system is implicated in the pathogenesis of a variety of diseases, including neurological disorders, which are pathologically characterized by the presence of ubiquitin-positive protein aggregates. A direct correlation between intact neuronal functioning and the UPS is exemplified by a range of transgenic mouse models wherein mutations in components of the UPS lead to a neurodegenerative or neurological phenotype. These models have been proven useful in determining the role of the UPS in the nervous system in health and disease. Furthermore, recently developed in vivo models harboring reporter systems to measure UPS activity could also substantially contribute to understanding the effect of neurodegeneration on UPS function. The role of the UPS in neurodegeneration in vivo is reviewed by discussing the currently available murine models showing a neurological phenotype induced by genetic manipulation of the UPS.