작품은 국립 보건원에서 보조금 1P01DA028555 - 01A1, 2R01 NS034239, 2R37 NS36126, P01 NS31492, P20RR 15635, P01MH64570 및 P01 NS43985 (HEG)을 지원했다. 저자는 원고와 뛰어난 그래픽과 문학 지원 중요한 읽기위한 양 로빈 테일러 감사합니다. 우리는 그의 축축한 밀링 전문 스티브 Grzelak 감사하고 싶습니다. 우리는 또한 검사 및 전송 전자 현미경 이미지를 공급 네브라스카 - 링컨 전자 현미경의 핵심 시설의 대학의 박사 한 첸 감사하고 싶습니다. 마지막으로, 우리는 살아 공촛점 현미경을 사용하여 그녀의 전문 메간 말쿼트 감사하고 싶습니다.

<ol>
	<li>Gendelman, H. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficient+isolation+and+propagation+of+human+immunodeficiency+virus+on+recombinant+colony-stimulating+factor+1-treated+monocytes.">Efficient isolation and propagation of human immunodeficiency virus on recombinant colony-stimulating factor 1-treated monocytes.</a> J Exp Med. 167 (4), 1428-1441 (1988).</li><li>Dou, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+a+macrophage-based+nanoparticle+platform+for+antiretroviral+drug+delivery.">Development of a macrophage-based nanoparticle platform for antiretroviral drug delivery.</a> Blood. 108 (8), 2827-2835 (2006).</li><li>Dou, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laboratory+investigations+for+the+morphologic,+pharmacokinetic,+and+anti-retroviral+properties+of+indinavir+nanoparticles+in+human+monocyte-derived+macrophages.">Laboratory investigations for the morphologic, pharmacokinetic, and anti-retroviral properties of indinavir nanoparticles in human monocyte-derived macrophages.</a> Virology. 358, 148-158 (2007).</li><li>Dou, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Macrophage+delivery+of+nanoformulated+antiretroviral+drug+to+the+brain+in+a+murine+model+of+neuroAIDS.">Macrophage delivery of nanoformulated antiretroviral drug to the brain in a murine model of neuroAIDS.</a> J Immunol. 183 (1), 661-669 (2009).</li><li>Nowacek, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoformulated+Antiretroviral+Drug+Combinations+Extend+Drug+Release+and+Antiretroviral+Responses+in+HIV-1-Infected+Macrophages:+Implications+for+NeuroAIDS+Therapeutics.">Nanoformulated Antiretroviral Drug Combinations Extend Drug Release and Antiretroviral Responses in HIV-1-Infected Macrophages: Implications for NeuroAIDS Therapeutics.</a> J Neuroimmune Pharmacol. , (2010).</li><li>Nowacek, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NanoART+synthesis,+characterization,+uptake,+release+and+toxicology+for+human+monocyte-macrophage+drug+delivery.">NanoART synthesis, characterization, uptake, release and toxicology for human monocyte-macrophage drug delivery.</a> Nanomed. 4 (8), 903-917 (2009).</li><li>Kalter, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+HIV+replication+in+macrophage+colony-stimulating+factor-treated+monocytes.">Enhanced HIV replication in macrophage colony-stimulating factor-treated monocytes.</a> J Immunol. 146 (1), 298-306 (1991).</li></ol>

관심 없음 충돌 선언하지 않습니다.

NanoART는 HIV - 1 치료제를 개선하기 위해 설계되었습니다. 우리는 임상 사용하기 위해 이러한 기술을 개발하는 첫 단계로서 nanoformulated 약물 개발을위한 실험실 테스트 시스템으로 단핵구 - 대식 세포 기반 약물 전달 시스템을 제안합니다. 우리의 과거 작품은 시스템이 그러한 응용 프로그램 5,6를위한 실용적입니다 것으로 나타났습니다. 현재 보고서에서, 우리는 일반적으로 사용되는 HIV - 1 치료 (; ritonovir, RTV, efavirenz, EFV 및 atazanavir, ATV indinavir, IDV)의 NP를 합성하는 방법을 그림 있습니다. 이것은 서부 유럽 표준시 밀링, 균질화 및 ultrasonication에 의해 달성되었다. 중요한 것은, 우리의 접근 방법은 크기, 모양 및 충전 5,6 등 nanoART 물리적 특성의 변경을 허용합니다. 계면 활성제의주의를 선택함으로써, 하나는 매우 긍정으로 중립에 매우 부정에서 입자의 요금을 제어할 수 있습니다. 처리 시간과 강도를 변경하여, 하나는 크기를 제어하고 ≥ 3 μm의 한 ≤ 200 nm의 작은 또는 큰 입자를 렌더링 수 있습니다. 입자의 모양도 있지만, 다른 물리적 특성을보다 낮은 범위를 조절할 수 있습니다. 멀티 양면 기하학적 형태가 서부 유럽 표준시 밀링 또는 균질 통해 생산 수 있지만 예를 들어, 구형 입자는 고분자와 sonication을 통해 만들 수 있습니다. 습식 밀링 및 균질 다르게 모양의 입자를 생산하지만,이 프로세스는 분류 방법에 의존하고 직접에 대해 제어할 수 없습니다. 제조 방법의 이러한 특성은 연구가 쉽게 정확한 설계 사양에 nanoART 생산 설정할 수 있습니다. nanoART의 물리 특성들이 안정성 모두에 강력한 영향을 어떻게 그들이 세포와 상호 작용 때문에 이것은 극히 중요합니다. 예를 들어, 크기가 약 1 μm의 아르 nanoART은 강한 긍정적인 요금을 가지고 있고, multisided 기하학적인 형태가 더 빠르게 한 번 세포 내부에는 더 안정 macrophages, 의해 촬영, 그리​​고 오랜 기간 동안 출시되는 것으로 나타났습니다 시간 6. 테스트 nanoART하는 방법은 최대 이동하는 입자 &#39;능력의 간단한 검사를 허용하는 개발 보존하고, 바이러스 복제를 억제하기 위해 용량 이외에, macrophages, HIV - 1의 원칙 대상 세포 중 하나에 의해 발표되었습니다. 이것은 하나가 쉽게 실적을 기반으로 nanoART 차별화 인간 사용 nanoART을 개발의 효율성과 속도를 모두 증가, 따라서, 생체내 모델의 성공의 가장 좋은 기회를 이들을 확인하실 수 있습니다. 이것은 마우스 골수 macrophages가 nanoART와 전 생체내 장전해서 adoptively 인간화 HIV - 1 감염 생쥐의 중추 신경계를 포함한 활성 감염의 사이트에 여행, 및 릴리스 약물 최대를 위해 할 수 이주로 정맥 주사에 의해 양도 때 표시되었습니다 바이러스성 복제에게 2-4 억제합니다. 또한, 이러한 nanoART로드 세포는 또한 효과적으로 플라즈마에서 바이러스에 감염된 세포의 숫자를 줄일 수 있었다뿐 아니라 CD4 + 세포 2 보호 등 임파선, 비장, 간, 폐. 이러한 연구는 예비 있지만, 셀 중재 nanoART 전달 시스템은 혈액과 감염된 조직 모두에게 약물의 임상 상당한 금액을 배포할 수있다는 것을 보여준다. 때문에 HIV - 1 감염 마우스 연구에서 고무적인 예비 결과, 우리는 현재 추가 임상 사용을위한 nanoART의 잠재력을 테스트하는 유인원 단백질 결핍 바이러스에 감염된 짧은 꼬리 원숭이 모델을 개발하고 있습니다.

테이블에서 사용되는 약어 : IDV : indinavir; RTV : ritonavir, ATV : atazanavir, EFV : effavirenz, PVA : polyvinylalcohol, SDS 나트륨 dodecyl 황산, P188 : poloxamer 188 (또한 Pluronic F68 칭했다), MPEG 2000 DSPE : 메틸 폴리 (에틸렌 - 글리콜) 1,2 - distearoyl - phosphatidyl - ethanolamine; DOTAP : (1 oleoyl - 2 - [6 - [(7 - 니트로 - 21 ,3 - benzoxadiazol - 4 - yl) 아미노] hexanoyl] - 3 - trimethylammonium 프로판), PLGA : 폴리 (락트 - 공동 glycolic 수 소산​​); CTAB : cetyltrimethyl 암모늄 브로마이드, DMEM : Dulbecco의 수정된 이글 중간; MCSF : 대식 세포 콜로니 - 자극 인자, HPLC : 고압 액체 크로마 토그래피, HIV - 1 : 인간 면역 결핍 바이러스 -1

methods development for blood borne macrophage carriage of nanoformulated antiretroviral drugs

antiretroviral (ART) 약물에 대한 약물 toxicities을 줄이기 위해 또한 제공하면서 Nanoformulated 약물 pharmacodynamics와 생체 이용률을 향상시킬 수 있습니다. 이를 위해, 저희 연구실 예술의 regimens을 단순화하는 nanomedicine의 원칙을 적용하고 준수 및 약물 pharmacokinetics을 개선하면서 같은 toxicities을 줄일 수 있습니다. 제조 nanoformulated ART (nanoART)에 대한 간단하고 신뢰할 수있는 방법이 표시됩니다. 순수한 약물의 입자 계면 활성제의 지질 코팅의 얇은 층에 의해 캡슐과 서부 유럽 표준시 밀링 또는 고압 균질에 의해 작은 것들로 큰 약물 결정 fractionating에 의해 생산됩니다. 다른 방법으로 무료로 약물은 고분자의 비말에서 일시 중지되었습니다. 본, 마약은 개별 nanosized 방울이 형성되기 전까지는 다음 ultrasonication에 의해 흥분 폴리머 내에 해산합니다. 동적 광 산란 및 현미경 검사는 입자 (입자 크기, 요금과 모양)의 물리적 특성을 특징. 그들의 생물 학적 속성 (세포 이해 및 유지, 세포 독성 및 antiretroviral 효능)이 인간 단핵구 파생 macrophages (MDM)로 결정됩니다. MDM은 정화를위한 원심 elutriation를 사용 leukopacks에서 분리된 인간의 말초 혈액 monocytes에서 파생됩니다. 이러한 혈액을 매개로 macrophages 인간 면역 결핍 바이러스에 nanoART 배포 (HIV)에 감염된 장기 세포 전송기로 사용할 수 있습니다. HIV - 1 치료에 대한 임상 nanoparticles로 사용할 수 antiretroviral 약물의 repackaging하는 준수를 개선하고 적극적으로 질병 결과에 영향을 미칠 수있는 것을 우리는 멋부리다.

Watch this Scientific Journal Video about Methods Development for Blood Borne Macrophage Carriage of Nanoformulated Antiretroviral Drugs at JoVE.com

Methods Development for Blood Borne Macrophage Carriage of Nanoformulated Antiretroviral Drugs

indinavir, ritonavir, efavirenz 및 atazanavir의 Nanoparticles 야한 밀링, 균질화 및 ultrasonication를 사용하여 제조되었습니다. 이러한 nanoformulations, 집합 대식 세포 기반 약물 전달을 평가 antiretroviral 치료 (nanoART)를 nanoformulated 칭했다. 단핵구 - 대식 세포의 파생 nanoART의 이해, 유지 및 지속적인 릴리스가 결정됩니다. 이러한 예비 연구 임상 사용 nanoART의 가능성을 제안한다.

Nanoformulated Antiretroviral 약물의 혈액 본 대식 세포 캐리지를위한 방법 개발

Nanoformulated drugs can improve pharmacodynamics and bioavailability while serving also to reduce drug toxicities for antiretroviral (ART) medicines. To ...

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Bioengineering

13.1K 조회수.  University of Nebraska Medical Center. indinavir, ritonavir, efavirenz 및 atazanavir의 Nanoparticles 야한 밀링, 균질화 및 ultrasonication를 사용하여 제조되었습니다. 이러한 nanoformulations, 집합 대식 세포 기반 약물 전달을 평가 antiretroviral 치료 (nanoART)를 nanoformulated 칭했다. 단핵구 - 대식 세포의 파생 nanoART의 이해, 유지 및 지속적인 릴리스가 결정됩니다. 이러한 예비 연구 임상 사용 nanoART의 가능성을 제안한다.

비디오: Methods Development for Blood Borne Macrophage Carriage of Nanoformulated Antiretroviral Drugs

Fluorescence detection methods for microfluidic droplet platforms

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that accompany system miniaturisation. Advantages include the ability to efficiently process nano- to femoto- liter volumes of sample, facile integration of functional components, an intrinsic predisposition towards large-scale multiplexing, enhanced analytical throughput, improved control and reduced instrumental footprints.1
In recent years much interest has focussed on the development of droplet-based (or segmented flow) microfluidic systems and their potential as platforms in high-throughput experimentation.2-4 Here water-in-oil emulsions are made to spontaneously form in microfluidic channels as a result of capillary instabilities between the two immiscible phases. Importantly, microdroplets of precisely defined volumes and compositions can be generated at frequencies of several kHz. Furthermore, by encapsulating reagents of interest within isolated compartments separated by a continuous immiscible phase, both sample cross-talk and dispersion (diffusion- and Taylor-based) can be eliminated, which leads to minimal cross-contamination and the ability to time analytical processes with great accuracy. Additionally, since there is no contact between the contents of the droplets and the channel walls (which are wetted by the continuous phase) absorption and loss of reagents on the channel walls is prevented.
Once droplets of this kind have been generated and processed, it is necessary to extract the required analytical information. In this respect the detection method of choice should be rapid, provide high-sensitivity and low limits of detection, be applicable to a range of molecular species, be non-destructive and be able to be integrated with microfluidic devices in a facile manner. To address this need we have developed a suite of experimental tools and protocols that enable the extraction of large amounts of photophysical information from small-volume environments, and are applicable to the analysis of a wide range of physical, chemical and biological parameters. Herein two examples of these methods are presented and applied to the detection of single cells and the mapping of mixing processes inside picoliter-volume droplets. We report the entire experimental process including microfluidic chip fabrication, the optical setup and the process of droplet generation and detection.

Droplet-based microfluidic platforms are promising candidates for high throughput experimentation since they are able to generate picoliter, self-compartmentalized vessels inexpensively at kHz rates. Through integration with fast, sensitive and high resolution fluorescence spectroscopic methods, the large amounts of information generated within these systems can be efficiently extracted, harnessed and utilized.

microfluidic 비말 플랫폼 형광 검출 방법

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that ...

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생체공학

Improved Method for the Preparation of a Human Cell-based, Contact Model of the Blood-Brain Barrier

The blood-brain barrier (BBB) comprises impermeable but adaptable brain capillaries which tightly control the brain environment. Failure of the BBB has been implied in the etiology of many brain pathologies, creating a need for development of human in vitro BBB models to assist in clinically-relevant research. Among the numerous BBB models thus far described, a static (without flow), contact BBB model, where astrocytes and brain endothelial cells (BECs) are cocultured on the opposite sides of a porous membrane, emerged as a simplified yet authentic system to simulate the BBB with high throughput screening capacity. Nevertheless the generation of such model presents few technical challenges. Here, we describe a protocol for preparation of a contact human BBB model utilizing a novel combination of primary human BECs and immortalized human astrocytes. Specifically, we detail an innovative method for cell-seeding on inverted inserts as well as specify insert staining techniques and exemplify how we use our model for BBB-related research.

Establishment of human models of the blood-brain barrier (BBB) can benefit research into brain conditions associated with BBB failure. We describe here an improved technique for preparation of a contact BBB model, which permits coculturing of human astrocytes and brain endothelial cells on the opposite sides of a porous membrane.

방법 혈액 - 뇌 장벽의 인간 세포 기반의 연락 모델의 준비를 위해 개선

The blood-brain barrier (BBB) comprises impermeable but adaptable brain capillaries which tightly control the brain environment. Failure of the BBB has ...

Methods for Characterizing the Co-development of Biofilm and Habitat Heterogeneity

Biofilms are surface-attached microbial communities that have complex structures and produce significant spatial heterogeneities. Biofilm development is strongly regulated by the surrounding flow and nutritional environment. Biofilm growth also increases the heterogeneity of the local microenvironment by generating complex flow fields and solute transport patterns. To investigate the development of heterogeneity in biofilms and interactions between biofilms and their local micro-habitat, we grew mono-species biofilms of Pseudomonas aeruginosa and dual-species biofilms of P. aeruginosa and Escherichia coli under nutritional gradients in a microfluidic flow cell. We provide detailed protocols for creating nutrient gradients within the flow cell and for growing and visualizing biofilm development under these conditions. We also present protocols for a series of optical methods to quantify spatial patterns in biofilm structure, flow distributions over biofilms, and mass transport around and within biofilm colonies. These methods support comprehensive investigations of the co-development of biofilm and habitat heterogeneity.

Biofilms have complex interactions with their surrounding environment. To comprehensively investigate biofilm-environment interactions, we present here a series of methods to create heterogeneous chemical environment for biofilm development, to quantify local flow velocity, and to analyze mass transport in and around biofilm colonies.

바이오 필름 및 서식지의 이질성의 공동 개발을 특성화하기위한 방법

Biofilms are surface-attached microbial communities that have complex structures and produce significant spatial heterogeneities. Biofilm development is ...

In Vitro Model of Physiological and Pathological Blood Flow with Application to Investigations of Vascular Cell Remodeling

Vascular disease is a common cause of death within the United States. Herein, we present a method to examine the contribution of flow dynamics towards vascular disease pathologies. Unhealthy arteries often present with wall stiffening, scarring, or partial stenosis which may all affect fluid flow rates, and the magnitude of pulsatile flow, or pulsatility index. Replication of various flow conditions is the result of tuning a flow pressure damping chamber downstream of a blood pump. Introduction of air within a closed flow system allows for a compressible medium to absorb pulsatile pressure from the pump, and therefore vary the pulsatility index. The method described herein is simply reproduced, with highly controllable input, and easily measurable results. Some limitations are recreation of the complex physiological pulse waveform, which is only approximated by the system. Endothelial cells, smooth muscle cells, and fibroblasts are affected by the blood flow through the artery. The dynamic component of blood flow is determined by the cardiac output and arterial wall compliance. Vascular cell mechano-transduction of flow dynamics may trigger cytokine release and cross-talk between cell types within the artery. Co-culture of vascular cells is a more accurate picture reflecting cell-cell interaction on the blood vessel wall and vascular response to mechanical signaling. Contribution of flow dynamics, including the cell response to the dynamic and mean (or steady) components of flow, is therefore an important metric in determining disease pathology and treatment efficacy. Through introducing an in vitro co-culture model and pressure damping downstream of blood pump which produces simulated cardiac output, various arterial disease pathologies may be investigated.

In Vitro Model of Physiological and Pathological Blood Flow with Application to Investigations of Vascular Cell Remodeling

This protocol replicates physiological or pathological blood flow in vitro to aid in determining cell response in disease pathologies. By introducing a pressure damping chamber downstream of a blood pump, blood flow across the vasculature can be recapitulated and imposed on a monolayer of vascular endothelium or a mimetic co-culture.

혈관 세포 리모델링의 조사에 생리의 체외 모델 및 응용 프로그램과 함께 병리학 혈액 흐름

Vascular disease is a common cause of death within the United States. Herein, we present a method to examine the contribution of flow dynamics towards ...

Fully Automated Centrifugal Microfluidic Device for Ultrasensitive Protein Detection from Whole Blood

Enzyme-linked immunosorbent assay (ELISA) is a promising method to detect small amount of proteins in biological samples. The devices providing a platform for reduced sample volume and assay time as well as full automation are required for potential use in point-of-care-diagnostics. Recently, we have demonstrated ultrasensitive detection of serum proteins, C-reactive protein (CRP) and cardiac troponin I (cTnI), utilizing a lab-on-a-disc composed of TiO2 nanofibrous (NF) mats. It showed a large dynamic range with femto molar (fM) detection sensitivity, from a small volume of whole blood in 30 min. The device consists of several components for blood separation, metering, mixing, and washing that are automated for improved sensitivity from low sample volumes. Here, in the video demonstration, we show the experimental protocols and know-how for the fabrication of NFs as well as the disc, their integration and the operation in the following order: processes for preparing TiO2 NF mat; transfer-printing of TiO2 NF mat onto the disc; surface modification for immune-reactions, disc assembly and operation; on-disc detection and representative results for immunoassay. Use of this device enables multiplexed analysis with minimal consumption of samples and reagents. Given the advantages, the device should find use in a wide variety of applications, and prove beneficial in facilitating the analysis of low abundant proteins.

This protocol demonstrates how to achieve femto molar detection sensitivity of proteins in 10 &#181;L of whole blood within 30 min. This can be achieved by using electrospun nanofibrous mats integrated in a lab-on-a-disc, which offers high surface area as well as effective mixing and washing for enhanced signal-to-noise ratio.

전혈에서 고감도 단백질 검출을위한 완전 자동화 된 원심 미세 유체 장치

Enzyme-linked immunosorbent assay (ELISA) is a promising method to detect small amount of proteins in biological samples. The devices providing a platform ...

Development of a Multicellular Three-dimensional Organotypic Model of the Human Intestinal Mucosa Grown Under Microgravity

Because cells growing in a three-dimensional (3-D) environment have the potential to bridge many gaps of cell cultivation in 2-D environments (e.g., flasks or dishes). In fact, it is widely recognized that cells grown in flasks or dishes tend to de-differentiate and lose specialized features of the tissues from which they were derived. Currently, there are mainly two types of 3-D culture systems where the cells are seeded into scaffolds mimicking the native extracellular matrix (ECM): (a) static models and (b) models using bioreactors. The first breakthrough was the static 3-D models. 3-D models using bioreactors such as the rotating-wall-vessel (RWV) bioreactors are a more recent development. The original concept of the RWV bioreactors was developed at NASA&#39;s Johnson Space Center in the early 1990s and is believed to overcome the limitations of static models such as the development of hypoxic, necrotic cores. The RWV bioreactors might circumvent this problem by providing fluid dynamics that allow the efficient diffusion of nutrients and oxygen. These bioreactors consist of a rotator base that serves to support and rotate two different formats of culture vessels that differ by their aeration source type: (1) Slow Turning Lateral Vessels (STLVs) with a co-axial oxygenator in the center, or (2) High Aspect Ratio Vessels (HARVs) with oxygenation via a flat, silicone rubber gas transfer membrane. These vessels allow efficient gas transfer while avoiding bubble formation and consequent turbulence. These conditions result in laminar flow and minimal shear force that models reduced gravity (microgravity) inside the culture vessel. Here we describe the development of a multicellular 3-D organotypic model of the human intestinal mucosa composed of an intestinal epithelial cell line and primary human lymphocytes, endothelial cells and fibroblasts cultured under microgravity provided by the RWV bioreactor.

Cells growing in a three-dimensional (3-D) environment represent a marked improvement over cell cultivation in 2-D environments (e.g., flasks or dishes). Here we describe the development of a multicellular 3-D organotypic model of the human intestinal mucosa cultured under microgravity provided by rotating-wall-vessel (RWV) bioreactors.

인간의 장 점막의 다세포 3 차원의 Organotypic 모델의 개발은 미세 중력 하에서 성장

Because cells growing in a three-dimensional (3-D) environment have the potential to bridge many gaps of cell cultivation in 2-D environments (e.g., ...

In Vitro Permeation of FITC-loaded Ferritins Across a Rat Blood-brain Barrier: a Model to Study the Delivery of Nanoformulated Molecules

Brain microvascular endothelial cells, supported by pericytes and astrocytes endfeet, are responsible for the low permeation of large hydrosoluble drugs through the blood-brain barrier (BBB), causing difficulties for effective pharmacological therapies. In recent years, different strategies for promoting brain targeting have aimed to improve drug delivery and activity at this site, including innovative nanosystems for drug delivery across the BBB. In this context, an in vitro approach based on a simplified cellular model of the BBB provides a useful tool to investigate the effect of nanoformulations on the trans-BBB permeation of molecules. This study describes the development of a double-layer BBB, consisting of co-cultured commercially available primary rat brain microvascular endothelial cells and astrocytes. A multiparametric approach for the validation of the model, based on the measurement of the transendothelial electrical resistance and the apparent permeability of a high molecular weight dextran, is also described. As proof of concept for the employment of this BBB model to study the effect of different nanoformulations on the translocation of fluorescent molecules across the barrier, we describe the use of fluorescein isothiocyanate (FITC), loaded into ferritin nanoparticles. The ability of ferritins to improve the trans-BBB permeation of FITC was demonstrated by flux measurements and confocal microscopy analyses. The results suggest this is a useful system for validating nanosystems for delivery of drugs across the BBB.

In Vitro Permeation of FITC-loaded Ferritins Across a Rat Blood-brain Barrier: a Model to Study the Delivery of Nanoformulated Molecules

A method to establish an in vitro model of blood-brain barrier based on a co-culture of rat brain microvascular endothelial cells and astrocytes is described and validated. This system proved to be a valid tool to study the effect of nanoformulation on the trans-barrier permeation of fluorescent molecules.

쥐 뇌 혈관 벽의 맞은 편에 FITC로드 Ferritins의 생체 외 투과 년 : 모델 Nanoformulated 분자의 배달을 공부하기

Brain microvascular endothelial cells, supported by pericytes and astrocytes endfeet, are responsible for the low permeation of large hydrosoluble drugs ...

Two Methods for Decellularization of Plant Tissues for Tissue Engineering Applications

The autologous, synthetic, and animal-derived grafts currently used as scaffolds for tissue replacement have limitations due to low availability, poor biocompatibility, and cost. Plant tissues have favorable characteristics that make them uniquely suited for use as scaffolds, such as high surface area, excellent water transport and retention, interconnected porosity, preexisting vascular networks, and a wide range of mechanical properties. Two successful methods of plant decellularization for tissue engineering applications are described here. The first method is based on detergent baths to remove cellular matter, which is similar to previously established methods used to clear mammalian tissues. The second is a detergent-free method adapted from a protocol that isolates leaf vasculature and involves the use of a heated bleach and salt bath to clear the leaves and stems. Both methods yield scaffolds with comparable mechanical properties and low cellular metabolic impact, thus allowing the user to select the protocol which better suits their intended application.

Here we present, and contrast two protocols used to decellularize plant tissues: a detergent-based approach and a detergent-free approach. Both methods leave behind the extracellular matrix of the plant tissues used, which can then be utilized as scaffolds for tissue engineering applications.

Decellularization 식물 조직의 조직 공학 응용 프로그램에 대 한 두 가지 방법

The autologous, synthetic, and animal-derived grafts currently used as scaffolds for tissue replacement have limitations due to low availability, poor ...

Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial bypass. Characterization of the growth dynamics and remodeling process of degradable polymer scaffold-based tissue-engineered blood vessels (TEBVs) with pulsatile stimulation is crucial for vascular tissue engineering. Optical imaging techniques stand out as powerful tools for monitoring vascularization of engineered tissue enabling high-resolution imaging in real-time culture. This paper demonstrates a nondestructive and fast real-time imaging strategy to monitor the growth and remodeling of TEBVs in long-term culture by using optical coherence tomography (OCT). Geometric morphology is evaluated, including vascular remodeling process, wall thickness, and comparison of TEBV thickness in different culture time points and presence of pulsatile stimulation. Finally, OCT provides practical possibilities for real-time observation of the degradation of polymer in the reconstructing tissues under pulsatile stimulation or not and in each vessel segment, by compared with the assessment of polymer degradation using scanning electron microscopic(SEM) and polarized microscope.

A step by step protocol for nondestructive and long-period monitoring the process of vascular remodeling and scaffold degradation in real-time culture of biodegradable polymeric scaffold-based tissue-engineered blood vessels with pulsatile stimulation using optical coherence tomography is described here.

광학 일관성 단층 촬영을 사용 하 여 분해 비 계 기반 조직 설계 혈관 개발의 비파괴 모니터링

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial ...

Intra-cardiac Side-Firing Light Catheter for Monitoring Cellular Metabolism using Transmural Absorbance Spectroscopy of Perfused Mammalian Hearts

Absorbance spectroscopy of cardiac muscle provides non-destructive assessment of cytosolic and mitochondrial oxygenation via myoglobin and cytochrome absorbance respectively. In addition, numerous aspects of the mitochondrial metabolic status such as membrane potential and substrate entry can also be estimated. To perform cardiac wall transmission optical spectroscopy, a commercially available side-firing optical fiber catheter is placed in the left ventricle of the isolated perfused heart as a light source. Light passing through the heart wall is collected with an external optical fiber to perform optical spectroscopy of the heart in near real- time. The transmission approach avoids numerous surface scattering interference occurring in widely used reflection approaches. Changes in transmural absorbance spectra were deconvolved using a library of chromophore reference spectra, providing quantitative measures of all the known cardiac chromophores simultaneously. This spectral deconvolution approach eliminated intrinsic errors that may result from using common dual wavelength methods applied to overlapping absorbance spectra, as well as provided a quantitative evaluation of the goodness of fit. A custom program was designed for data acquisition and analysis, which permitted the investigator to monitor the metabolic state of the preparation during the experiment. These relatively simple additions to the standard heart perfusion system provide a unique insight into the metabolic state of the heart wall in addition to conventional measures of contraction, perfusion, and substrate/oxygen extraction.

Here we introduce a method for using an intra-ventricle optical catheter in perfused hearts to perform absorbance spectroscopy across the heart wall. The data obtained provides robust information on tissue oxygen tension as well as substrate utilization and membrane potential simultaneously with cardiac performance measures in this ubiquitous preparation.

교란 성 포유류 심장의 경부 흡광도 분광학을 사용하여 세포 대사를 모니터링하기위한 심장 내 측 발사 라이트 카테터

Absorbance spectroscopy of cardiac muscle provides non-destructive assessment of cytosolic and mitochondrial oxygenation via myoglobin and cytochrome ...