Massively-Parallelized, Deterministic Mechanoporation for Intracellular Delivery

Microfluidic intracellular delivery approaches based on plasma membrane poration have shown promise for addressing the limitations of conventional cellular engineering techniques in a wide range of applications in biology and medicine. However, the inherent stochasticity of the poration process in many of these approaches often results in a trade-off between delivery efficiency and cellular viability, thus potentially limiting their utility. Herein, we present a novel microfluidic device concept that mitigates this trade-off by providing opportunity for deterministic mechanoporation (DMP) of cells en masse. This is achieved by the impingement of each cell upon a single needle-like penetrator during aspiration-based capture, followed by diffusive influx of exogenous cargo through the resulting membrane pore, once the cells are released by reversal of flow. Massive parallelization enables high throughput operation, while single-site poration allows for delivery of small and large-molecule cargos in difficult-to-transfect cells with efficiencies and viabilities that exceed both conventional and emerging transfection techniques. As such, DMP shows promise for advancing cellular engineering practice in general and engineered cell product manufacturing in particular

Efficient Dielectrophoretic Patterning of Embryonic Stem Cells in Energy Landscapes Defined by Hydrogel Geometries

In this study, we have developed an integrated microfluidic platform for actively patterning mammalian cells, where poly(ethylene glycol) (PEG) hydrogels play two important roles as a non-fouling layer and a dielectric structure. The developed system has an embedded array of PEG microwells fabricated on a planar indium tin oxide (ITO) electrode. Due to its dielectric properties, the PEG microwells define electrical energy landscapes, effectively forming positive dielectrophoresis (DEP) traps in a low-conductivity environment. Distribution of DEP forces on a model cell was first estimated by computationally solving quasi-electrostatic Maxwell’s equations, followed by an experimental demonstration of cell and particle patterning without an external flow. Furthermore, efficient patterning of mouse embryonic stem (mES) cells was successfully achieved in combination with an external flow. With a seeding density of 107 cells/mL and a flow rate of 3 μL/min, trapping of cells in the microwells was completed in tens of seconds after initiation of the DEP operation. Captured cells subsequently formed viable and homogeneous monolayer patterns. This simple approach could provide an efficient strategy for fabricating various cell microarrays for applications such as cell-based biosensors, drug discovery, and cell microenvironment studies

頭頸部領域および食道領域の各種症状に対する食道運動機能の関与について : High Resolution Manometry を用いた検討

頭頸部領域および食道領域における各種症状に対する食道運動機能の関与を検討した.2007年9月から2012年6月までに,咽喉頭異常感などの頭頸部領域の症状および嚥下困難感,胸痛,胸やけなどの食道領域に関連した症状を主訴に,当科を受診した261例(男性138例,女性123例,平均年齢56.8±17.1才)を対象とし,健康関連QOL(Health Related Quality of Life: HRQL)の測定と食道内圧検査を施行した.健康関連QOLの検討では,咽喉頭違和感,嚥下困難感,胸やけ等の各症状を訴えた患者で,身体的QOL,精神的QOL を表すPCS(physical component summary)あるいはMCS(mental component summary)が低下し,健常者と比較して有意にQOLの低下を認めた.食道内圧検査による食道運動機能異常は,全対象患者中62.0%に認めた.各症状別に食道運動機能障害の内訳を見ると,咽喉頭違和感ではIEM(ineffective esophageal motility)(31.8%),嚥下困難感は食道アカラシア(56.6%),喉のつかえ感は食道アカラシア(35.5%),胸やけはIEM(39.4%),胸痛は食道アカラシア(50.0%),噫気はIEM(50.0%)を最も多く認めた.咽喉頭違和感,嚥下困難感,喉のつかえ感,胸やけ,胸痛などの頭頚部および食道症状を有するものの,器質的疾患を認めない患者のQOLは障害されており,その病態の一つとして食道運動機能異常の存在を念頭に置き,診療にあたることが重要である.There are many cases of patients presenting at the hospital complaining of dysphagia or chest pain. At that time organic diseases are not always detected in these patients;previously several studies showed esophageal motility dysfunction was one of the important factors in the development of these symptoms. However, there have been few reports investigating the relationship between esophageal motility dysfunction and these symptoms. Between September 2007 and June 2012, 216 patients (138 men,123 women;mean age 56.8±17.1 years old) were admitted complaining of the following symptoms:dysphagia, globus sensation, or heart burn and non-cardiac chest pain,but without any organic diseases related symptoms. They all underwent high-resolution esophageal manometry (HRM) and completed a questionnaire regarding their health-related quality of life (HRQOL). The aim of this study was to evaluate the relationship between these symptoms and esophageal motility dysfunction and QOL. In our series, 62% of patients with non-obstructive dysphagia, globus sensation, heartburn and unexplained (non-cardiac) chest pain have an esophageal motility disorder (Achalasia 31.5%, IEM 26.4%, Diffuse esophageal spasm 4.6%;nutcracker esophagus 2.8% and Impaired ECJ relaxation 0.5%). SF-8 score of PCS(Physical Component Summary) in all of the symptoms were significantly lower in comparison with the matched healthy control. SF-8 score of MCS(Mental Component Summary) with globus sensation and esophageal dysphagia were significantly lower in comparison with the matched healthy control. The patients complaining of several symptoms located in the pharynx or esophagus have impaired quality of life and esophageal motility dysfunctions.It is important to consider these issues while examining these patients