Transport Of Various Substances Research Articles

Blood–brain barrier microfluidic chips and their applications

As a natural dynamic barrier separating blood from brain parenchyma, the blood–brain barrier (BBB) is mainly composed of brain microvascular endothelial cells (BMECs), pericytes, astrocytes, and a variety of neurons. The BBB regulates the highly selective transport of various substances between the brain and blood and maintains the stability of the central nervous system (CNS). Owing to this tight control, the BBB represents a formidable challenge for the delivery of drugs and other exogenous compounds into the CNS, which has bottlenecked the development of many drugs for CNS diseases. Therefore, efficient and precise in vitro models of the BBB are needed to assess the efficacy and toxicity of drugs targeting the CNS to inform drug design and to improve the success rate of agents that enter clinical evaluation. In vitro BBB models have rapidly advanced from the early two-dimensional (2D) static models to the current three-dimensional (3D) dynamic microfluidic chips. Although the commonly used, static, in vitro BBB models are simple to construct and TEER values are convenient to detect, the static models do not provide an ideal (i.e., accurate) BBB environment, since they lack the correct physiological size/scale and hemodynamic shear stress, both of which play substantial roles in promoting and maintaining EC differentiation into a specific BBB phenotype. Compared with traditional static models, 3D microfluidic models thus enable cells to react in a manner more closely resembling in vivo behavior by simulating a microenvironment with more natural signal transduction. As a result, the dynamic 3D BBB model can more accurately recapitulate the structure and function of the human BBB. Here we summarize the recent progress in in vitro microfluidic BBB chips and their research applications as well as discuss the prospects and challenges for where the technology is headed.

Clinical Significance of Lipid Transport Function of ABC Transporters in the Innate Immune System.

ABC transporters are a large family of proteins that transport a variety of substrates across cell plasma membranes. Because of this, they are involved in many physiological processes. It is of interest to note that many ABC transporters are involved in the transport of various lipids. In addition, this function may be related to the innate immune system. The evidence that ABC transporters are involved in the regulation of the innate immune system through the transport of various substances greatly enhances the understanding of their clinical significance. ABC transporters are involved in the cellular homeostasis of cholesterol as well as in the regulation of its content in lipid rafts. Through these mechanisms, they can regulate the function of membrane proteins, including receptors of the innate immune system. By regulating lipid transport, some members of ABC transporters are involved in phagocytosis. In addition, ABC transporters are involved in the transport of lipopolysaccharide, lipid mediators of inflammation, and perform other functions in the innate immune system.

Structural and functional investigation of ABC transporter STE6-2p from Pichia pastoris reveals unexpected interaction with sterol molecules

Adenosine triphosphate (ATP)-binding cassette (ABC) transporters are multidomain transmembrane proteins, which facilitate the transport of various substances across cell membranes using energy derived from ATP hydrolysis. They are important drug targets since they mediate decreased drug susceptibility during pharmacological treatments. For the methylotrophic yeast Pichia pastoris, a model organism that is a widely used host for protein expression, the role and function of its ABC transporters is unexplored. In this work, we investigated the Pichia ABC-B transporter STE6-2p. Functional investigations revealed that STE6-2p is capable of transporting rhodamines invivo and is active in the presence of verapamil and triazoles invitro. A phylogenetic analysis displays homology among multidrug resistance (MDR) transporters from pathogenic fungi to human ABC-B transporters. Further, we present high-resolution single-particle electron cryomicroscopy structures of an ABC transporter from P. pastoris in the apo conformation (3.1 Å) and in complex with verapamil and adenylyl imidodiphosphate (AMP-PNP) (3.2 Å). An unknown density between transmembrane helices 4, 5, and 6 in both structures suggests the presence of a sterol-binding site of unknown function.

Comprehensive understanding of multiple actions of anticancer drug tamoxifen in isolated mitochondria

Tamoxifen has been widely used in the treatment of estrogen receptor (ER)-positive breast cancer, whereas it also exhibits ER-independent anticancer effects in various cancer cell types. As one of the convincing mechanisms underlying the ER-independent effects, induction of apoptosis through mitochondrial dysfunction has been advocated. However, the mechanism of action of tamoxifen even at the isolated mitochondrial level is not fully understood and remains controversial. Here, we attempted to comprehensively understand tamoxifen's multiple actions in isolated rat liver mitochondria through not only revisiting the actions hitherto reported but also conducting originally designed experiments. Using submitochondrial particles, we found that tamoxifen has potential as an inhibitor of both respiratory complex I and ATP synthase. However, these inhibitory effects were not elicited in intact mitochondria, likely because penetration of tamoxifen across the inner mitochondrial membrane is highly restricted owing to its localized positive charge (-N+H(CH3)2). This restricted penetration may also explain why tamoxifen is unable to function as a protonophore-type uncoupler in mitochondria. Moreover, tamoxifen suppressed opening of the mitochondrial permeability transition pore induced by Ca2+ overload through enhancing phosphate uptake into the matrix. The photoaffinity labeling experiments using a photolabile tamoxifen derivative (pTAM1) indicated that pTAM1 specifically binds to voltage-dependent anion channels (VDACs) 1 and 3, which regulate transport of various substances into mitochondria. The binding of tamoxifen to VDAC1 and/or VDAC3 could be responsible for the enhancement of phosphate uptake. Taking all the results together, we consider the principal impairment of mitochondrial functions caused by tamoxifen.

The rostral epidural rete mirabile of the llama as a place of retrograde transport of various substances – anatomical basics

Abstract The aim of this study was to analyse the structure of the rostral epidural rete mirabile in the llama. Some specimens were prepared by injecting stained chemically cured acrylic into the bilateral common carotid arteries. After about 1 month received vascular corrosion casts on the bone scaffold. Some specimens made using red and blue latex introduced into the bilateral common carotid arteries and the bilateral external jugular vein. The rostral epidural rete mirabile is a well-developed, bilateral structure composed of numerous arteries, which are multiply anastomosed with each other. The cranial section of the rete is asymmetrical. Its lateral part is much better developed, because there are rostral branches to the rostral epidural rete mirabile at this point. The arterial vessels are not accompanied by homonymous veins. However, the arteries of the rostral epidural rete mirabile are accompanied by venous vessels of the cavernous sinus. That rete plays an important role in selective brain cooling, the conservation of body water, and retrograde transport of neurotransmitters. CO, GnRH, beta-endorphin, progesterone, testosterone, oxytocin, LHRH and dopamine diffuse from the venous blood of the cavernous sinus to the arterial blood of the rostral epidural rete mirabile.

Rhodochrosite Oxidation by Dissolved Oxygen and the Formation of Mn Oxide Product: The Impact of Goethite as a Foreign Solid Substrate.

Rhodochrosite conversion to Mn (oxyhydr)oxides significantly affects the fate and transport of various substances in the environment. We examined rhodochrosite oxidation by dissolved oxygen and the oxidation product formation with an emphasis on the effects of goethite substrate. Without goethite, rhodochrosite oxidation was slow as no detectable change was observed for 28 d with microscopic and spectroscopic analyses, except a minor change in X-ray diffraction. Interestingly, by contrast, it was greatly accelerated and completed in 7 d in the presence of goethite, resulting in the heteroepitaxial growth of groutite (α-MnOOH)-like Mn oxides on the goethite (α-FeOOH) tip surfaces. The formation of this secondary Mn oxide likely induced the acidification of the microenvironment in the vicinity of rhodochrosite particles and thereby promoted their dissolution. Subsequently, their oxidative conversion to the Mn oxide was expedited by the surface catalyzed Mn(II) oxidation on the goethite tip. Our results revealed that goethite as a foreign substrate imparts a decisive control on not only the rate but also the type of the reaction product of rhodochrosite oxidation. This study presents a new insight into the geochemical roles of foreign particles on the dynamics of redox-sensitive solid phases in the environment.

Development of an Organ-on-a-Chip-Device for Study of Placental Pathologies.

The human placenta plays a key role in reproduction and serves as a major interface for maternofetal exchange of nutrients. Study of human placenta pathology presents a great experimental challenge because it is not easily accessible. In this paper, a 3D placenta-on-a-chip model is developed by bioengineering techniques to simulate the placental interface between maternal and fetal blood in vitro. In this model, trophoblasts cells and human umbilical vein endothelial cells are cultured on the opposite sides of a porous polycarbonate membrane, which is sandwiched between two microfluidic channels. Glucose diffusion across this barrier is analyzed under shear flow conditions. Meanwhile, a numerical model of the 3D placenta-on-a-chip model is developed. Numerical results of concentration distributions and the convection–diffusion mass transport is compared to the results obtained from the experiments for validation. Finally, effects of flow rate and membrane porosity on glucose diffusion across the placental barrier are studied using the validated numerical model. The placental model developed here provides a potentially helpful tool to study a variety of other processes at the maternal–fetal interface, for example, effects of drugs or infections like malaria on transport of various substances across the placental barrier.

Ultrastructural Changes Associated With the Enhanced Permeability of the Round Window Membrane Mediated by Ultrasound Microbubbles.

The round window membrane (RWM) is the most common entryway for local drug and gene delivery into the inner ear, but its permeability can change the treatment outcome. We previously demonstrated a feasible and highly efficient approach using ultrasound-aided microbubble (USMB) cavitation to enhance the permeability of the RWM. Here, we investigated the safety of USMB exposure and the association between temporal changes in RWM permeability and ultrastructure. Experimental guinea pigs were divided into two treatment groups: a control group receiving round window soaking (RWS) with MBs and treatment (USM) groups undergoing 3 (USM-3) or 5 (USM-5) consecutive USMB exposures (1 min/exposure) at an acoustic intensity of 3 W/cm2 and 1 MHz frequency. The trans-RWM delivery efficiency of biotin-fluorescein isothiocyanate conjugates, used as permeability tracers, revealed a greater than 7-fold higher delivery efficiency for the USM groups immediately after 3 or 5 exposures than for the RWS group. After 24 h, the delivery efficiency was 2.4-fold higher for the USM-3 group but was 6.6-fold higher for the USM-5 group (and 3.7-fold higher after 48 h), when compared to the RWS group. Scanning electron microscopy images of the RWM ultrastructure revealed USMB-induced sonoporation effects that could include the formation of heterogeneous pore-like openings with perforation diameters from 100 nm to several micrometers, disruption of the continuity of the outer epithelial surface layer, and loss of microvilli. These ultrastructural features were associated with differential permeability changes that depended on the USMB exposure course. Fourteen days after treatment, the pore-like openings had significantly decreased in number and the epithelial defects were healed either by cell expansion or by repair by newly migrated epithelial cells. The auditory brainstem response recordings of the animals following the 5-exposure USMB treatment indicated no deterioration in the hearing thresholds at a 2-month follow-up and no significant hair cell damage or apoptosis, based on scanning electron microscopy, surface preparations, and TUNEL assays. USMBs therefore appear to be safe and effective for inner ear drug delivery. The mechanism of enhanced permeability may involve a disruption of the continuity of the outer RWM epithelial layer, which controls transmembrane transport of various substances.

Involvement of plasma membrane H+‐ATPase in the ammonium‐nutrition response of barley roots

AbstractBarley (Hordeum vulgare L.) is a typical ammonium ()‐sensitive crop species and exhibits a futile‐ efflux from root cells. Plasma membrane (PM) H+‐ATPase is responsible for pumping H+ out of plant cells and providing the driving force for the transport of various substances. We hypothesized that PM H+‐ATPase is involved in this efflux process of barley roots. Barley plants were cultivated in hydroponic solution with 1.0 mM or 1.0 mM nitrate () as the sole nitrogen source. Plasma membrane vesicles were isolated from the root tips for analyses of the activity. Plasma membrane H+‐ATPase was tested by Western Blot. The transcription of PM H+‐ATPase genes was analyzed using qRT‐PCR. Proton and efflux from barley roots was analyzed by scanning ion‐selective electrode technique. The results show that root and shoot growth was repressed by nutrition when compared to nutrition. Plasma membrane H+‐ATPase activity of barley roots under nutrition was significantly higher than that under nutrition. Methyl‐ammonium (MeA), a non‐metabolizable analog of , had no significant effect on PM H+‐ATPase activity. The enhanced PM H+‐ATPase activity by nutrition was consistent with a higher abundance of PM H+‐ATPase protein and higher transcription levels of six PM H+‐ATPase genes. By using the pharmacological agents vanadate and fusicoccin, a close linkage was observed between H+ efflux and efflux from barley roots. Taken together, nutrition stimulates PM H+‐ATPase activity, and this stimulation is possibly triggered by assimilation in barley roots. efflux from barley roots is associated with PM H+‐ATPase activity which could provide the driving force for efflux. Ammonium efflux might be considered as a disposal strategy in barley roots which prevents overloading the root cytoplasm.

Hepatic effect of the sub-chronic exposure to mosquito coil smoke in albino mice

The hepatic effect of chronic inhalation of mosquito coil smoke in mice was investigated. Mosquito coil smoke is mosquito repellent incense made from dried pyrethrum powder. It is widely used in Africa, Asia and South America. Exposure to mosquito coil smoke may lead to coughing, sneezing, and inflammation of trachea, renal damage, weight loss, headache, itching and skin reaction. It decreases the protein biosynthetic activity of the liver. This could affect capacity of serum protein-mediated transport of various substances. Goldeer mosquito coil containing 0.03% Transfluthrin, manufactured in Kano, Nigeria and purchased in Zaria was used for this experiment. Twenty-four (24) male mice (20-40 g), were divided into four groups of six mice each and exposed for six weeks to mosquito coil smoke for 1, 2 and 3 hours in an inhalation chamber. The animals were humanely sacrificed and the their blood samples collected and analysed for the enzymes, Aspartate amino transferase (AST), Alanine aminotransferase (ALT) and Alkaline Phosphatase (ALP) as markers of Hepatic function. Also, their brain tissues were collected, homogenized and centrifuged and the supernatants assayed for oxidative stress biomarker, malondialdehyde (MDA) and antioxidant enzymes, superoxide dismutase (SOD) and Catalase (CAT). Data were analysed using one way ANOVA and expressed as Mean±SEM with value of P<0.05 considered statistically significant. The result showed that the mosquito coil smoke exposure in mice does not cause any significant change in liver enzymes, MDA and SOD. However, there were significant increases in CAT activity (43.83±1.85) and (41.67±1.36) as compared to control (38.50±1.00).

Solute carriers (SLCs) identified and characterized from kidney transcriptome of golden mahseer (Tor putitora) (Fam: Cyprinidae).

The solute carriers (SLC) are trans-membrane proteins, those regulate the transport of various substances (sugars, amino acids, nucleotides, inorganic cations/anions, metals, drugs etc.) across the cell membrane. There are more than 338 solute carriers (slc) reported in fishes that play crucial role in cellular influx and efflux. The study of solute carrier families may reveal many answers regarding the function of transporter genes in the species and their effect in the existing environment. Therefore, we performed RNA sequencing of kidney tissue of the golden mahseer (Tor putitora) using Illumina platform to identify the solute carrier families and characterized 24 putative functional genes under 15 solute carrier families. Out of 24 putative functional genes, 11 genes were differentially expressed in different tissues (head kidney, trunk kidney, spleen, liver, gill, muscle, intestine and brain) using qRT-PCR assay. The slc5a1, slc5a12, slc12a3, slc13a3, slc22a13 and slc26a6 were highly expressed in kidney. The slc15a2, slc25a47, slc33a1 and slc38a2 were highly expressed in brain and slc30a5 was over-expressed in gill. The unrooted phylogenetic trees of slc2, slc5, slc13 and slc33 were constructed using amino acid sequences of Homo sapiens, Salmo salar, Danio rerio, Cyprinus carpio and Tor putitora. It appears that all the putative solute carrier families are very much conserved in human and fish species including the present fish, golden mahseer. This study provides the first hand database of solute carrier families particularly transporter encoding proteins in the species.

How hormones influence composition and physiological function of the brain-blood barrier.

Hormones exert many actions in the brain. Their access and effects in the brain are regulated by the blood-brain barrier (BBB). Hormones as other substances may enter the brain and vice versa either by paracellular way requiring breaching tight junctions stitching the endothelial cells composing the BBB, or by passage through the cells (transcellular way). Hormones influence both ways through their receptors, both membrane and intracellular, present on/in the BBB. In the review the main examples are outlined how hormones influence the expression and function of proteins forming the tight junctions, as well as how they regulate expression and function of major protein transporters mediating transport of various substances including hormone themselves.

Transendothelial Transport and Its Role in Therapeutics.

Present review paper highlights role of BBB in endothelial transport of various substances into the brain. More specifically, permeability functions of BBB in transendothelial transport of various substances such as metabolic fuels, ethanol, amino acids, proteins, peptides, lipids, vitamins, neurotransmitters, monocarbxylic acids, gases, water, and minerals in the peripheral circulation and into the brain have been widely explained. In addition, roles of various receptors, ATP powered pumps, channels, and transporters in transport of vital molecules in maintenance of homeostasis and normal body functions have been described in detail. Major role of integral membrane proteins, carriers, or transporters in drug transport is highlighted. Both diffusion and carrier mediated transport mechanisms which facilitate molecular trafficking through transcellular route to maintain influx and outflux of important nutrients and metabolic substances are elucidated. Present review paper aims to emphasize role of important transport systems with their recent advancements in CNS protection mainly for providing a rapid clinical aid to patients. This review also suggests requirement of new well-designed therapeutic strategies mainly potential techniques, appropriate drug formulations, and new transport systems for quick, easy, and safe delivery of drugs across blood brain barrier to save the life of tumor and virus infected patients.

Effect of phytohormones on absorption and distribution of ions in salt-stressed bean plants

Bean plant seedlings grown in water culture were treated for 5 days either with NaCl or with 7-times concentrated nutrient solution (diminished water potential by 3-10<sup>3</sup> hPa in both cases). Control and stressed plants were treated for 24 hrs with zeatin and GA,. NaCl-stress reduced distinctly ion absorption rate (K, Ca and P). Zeatin and GA<sub>3</sub> promoted potassium uptake, but only in NaCI-treated plants. These hormones diminished Na accumulation in metabolically active organs but increased P- and Ca-content. In plants grown under both kind of stresses zeatin and GA<sub>3</sub> partially reestablished the ratio of the main mono- to divalent cations, which increased in the leaves and apical part of the stressed plants. ABA introduced into the nutrient solution caused inhibition of the ion uptake (K, Ca, Mg and P). similar to that caused by NaCl-stress. The above reported results seem to confirm the supposition, that hormones act as an important factor contributing to regulation of both uptake and distribution of ions. In this way growth substances may also participate in the regulation of transport of various substances (among others - assimilates) in the whole plant.

Michael William Blackburn Bradbury 1930–2013

Michael William Blackburn Bradbury 1930–2013

Prognostic value of ABCG2 in moderately and poorly differentiated intrahepatic cholangiocarcinoma

Intrahepatic cholangiocarcinoma (ICC) is a primary hepatic malignancy derived from cholangiocytes. The survival rate of ICC patients is very low, and conventional chemotherapy is not effective in prolonging long-term survival. Adenosine 5'-triphosphate (ATP)-binding cassette (ABC) transporters mediate the transport of various substances in several cellular processes. The expression of ABCB1, ABCC1 and ABCG2 has been implicated in multidrug resistance and poor prognosis in several types of cancer. The aim of this study was to examine their expression in normal cholangiocytes and ICC tissues. Immunohistochemistry was employed to evaluate the expression of these transporters in 60 cases of ICC with respect to clinicopathological features and patient outcome. The proportions of cases with loss of ABCB1, ABCC1 and ABCG2 expression were 93.3%, 68.3% and 50%, respectively. Only the loss of ABCG2 was related to a worse prognosis (P = 0.031), and was associated with lymph node involvement (P = 0.003) and higher tumour grade (P = 0.028). Furthermore, multivariate analysis showed that the loss of ABCG2 expression was an independent prognostic factor in patients with moderately or poorly differentiated ICC (P = 0.02). These results suggest that ABCG2 may be involved in cholangiocarcinogenesis; the loss of its expression may enhance tumour progression and contribute to aggressive growth of ICC.

Barrier functions and paracellular integrity in human cell culture models of the proximal respiratory unit

Airway epithelial cells provide a barrier to the translocation of inhaled materials. Tight (TJ) and adherens junctions (AJ) play a key role in maintaining barrier functions, and are responsible for the selective transport of various substances through the paracellular pathway. In this study we compared a bronchial cell line (16HBE14o-) and primary bronchial cells (HBEC), both cocultivated with the fibroblast cell line Wi-38, with respect to their structural differentiation and their reaction to cytokine stimulation. HBEC formed a pseudostratified epithelial layer and expressed TJ and AJ proteins after 2 weeks in coculture. Mucus-producing and ciliated cells were found within 24 days. Additionally, a beating activity of the ciliated HBEC (14–19 Hz) could be detected. 16HBE14o- in coculture showed a multilayered growth without differentiation to a pseudostratified airway epithelium. Simultaneous exposure to TNF-α- and IFN-γ-induced significant changes in barrier function and paracellular permeability in the cocultures of HBEC/Wi-38 but not in the 16HBE14o-/Wi-38. In summary, HBEC in coculture mimic the structure of native polarized bronchial epithelium showing basal, mucus-producing and ciliated cells. Our system provides an opportunity to examine the factors that influence barrier and mucociliary function of bronchial epithelium within a time frame of 3 weeks up to 3 months in an in vivo-like differentiated model.

Numerical modelling of multiphase flow and transport processes in landfills

Waste material in municipal landfills can be described as heterogeneous porous media, where flow and transport processes of gases and liquids are combined with local material degradation. This paper deals with the basic formulation of a multiphase flow and transport model applicable to the numerical analysis of coupled transport and reaction processes inside landfills. The transport model treats landfills within the framework of continuum mechanics, where flow and transport processes are described on a macroscopic level. The composition of organic and inorganic matter in the solid phase and its degradation are modelled on a microscopic scale. The degradation model captures the different reaction schemes of various microbial activities. Subsequently, transport and reaction processes have to be coupled, since emissions at the surface and from the drainage layer depend on the flow of leachate and gas, the transport of various substances and heat, and the biodegradation of organic matter. The theoretical considerations presented here are fundamental to the development of numerical models for the simulation of multiphase flow and transport processes inside landfills coupled with biochemical reactions and heat generation. The implicit modelling of leachate and gas flows including growth and decay of micro-organisms are innovative contributions to landfill modelling

Relationship between Microtubule Network Structure and Intracellular Transport in Cultured Endothelial Cells Affected by Shear Stress

Endothelial cells (ECs) that line the inner surface of blood vessels are barriers to the transport of various substances into or from vessel walls, and are continuously exposed to shear stress induced by blood flow in vivo. Shear stress affects the cytoskeleton (e.g., microtubules, microfilaments, intermediate filaments), and affects the transport of macromolecules. Here, the relationship between the microtubule network structure and this transport process for albumin uptake within cultured aortic endothelial cells affected by shear stress was studied. Based on fluorescent images of albumin uptake obtained by using confocal laser scanning microscopy (CLSM), both the microtubule network and albumin uptake in ECs were disrupted by colchicine and were affected by shear stress loading.

A forty-two year voyage through vision research.

This presentation is an overview of my involvement in vision research and the factors and individuals that influenced my career in this field over the last 42 years. It also summarizes my research interests and contributions in the areas of aqueous humor dynamics, transport of various substances across blood-aqueous barrier and in the lens. The metabolism and function of glutathione in the lens and the development of tissue culture of human lens epithelium as a model system to study its role in lens and cataract formation are reviewed.