Poly-l-Arginine Enhances Paracellular Permeability via Serine/Threonine Phosphorylation of ZO-1 and Tyrosine Dephosphorylation of Occludin in Rabbit Nasal Epithelium

Ohtake, Kazuo; Maeno, Takuya; Ueda, Hideo; Ogihara, Masahiko; Natsume, Hideshi; Morimoto, Yasunori

doi:10.1023/b:pham.0000003383.86238.d1

Cited by 62 publications

(53 citation statements)

References 33 publications

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“…In bovine retinal endothelial cells in vitro, tyrosine phosphorylation of ZO-1 is associated with increased permeability through the paracellular pathway (39). ZO-1 also has serine phosphorylation sites (40), and serine/threonine phosphorylation of ZO-1 is associated with increased paracellular permeability in rabbit nasal epithelium (41). The type of injury may also influence ZO-1 phosphorylation.…”

Section: Zo-1 and Diabetic Nephropathymentioning

confidence: 99%

ZΟ-1 Expression and Phosphorylation in Diabetic Nephropathy

et al. 2006

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Cellular mechanisms responsible for the loss of capillary wall permselectivity in diabetic nephropathy are not well characterized. ZO-1 is a junctional protein involved in the assembly and proper function of a number of tight junctions and is also expressed at the junction of podocytes with the slit diaphragm. We investigated the effect of diabetes and high glucose concentration on the expression of ZO-1 in animal models of both type 1 and 2 diabetes and in rat glomerular epithelial cells. In diabetic animals, immunohistochemistry and Western blotting showed decreased expression of ZO-1 in glomeruli. Immunogold electron microscopy revealed redistribution of ZO-1 from the podocyte membrane to the cytoplasm in the diabetic animals. Exposure of rat glomerular epithelial cells to high glucose resulted in a decrease in the intensity of ZO-1 staining and redistribution of ZO-1 from the membrane to the cytoplasm, changes that are attenuated by blockade of the angiotensin II type 1 receptor. ZO-1 protein expression and serine and tyrosine phosphorylation of ZO-1 were also decreased in cells exposed to high glucose. These findings suggest that alterations in the content and localization of ZO-1 may be relevant to the pathogenesis of proteinuria in diabetes. Diabetes 55:894 -900, 2006 D iabetic nephropathy is characterized by proteinuria and progressive fibrosis, resulting in a decline in kidney function in humans and experimental models of diabetes. Proteinuria is one of the most important prognostic risk factors for kidney disease progression (1). Cellular and molecular mechanisms underlying the loss of glomerular permselectivity in diabetic nephropathy and other proteinuric states are under intense investigation. Recent studies have identified mutations in the genes encoding podocyte structural proteins nephrin, podocin, CD2AP, neph-1, and ␣-actinin 4, which result in changes in glomerular permeability (2). ZO-1 ␣Ϫ, an 80 -amino acid truncated isoform of the 225-kD tight junction protein ZO-1, is highly expressed within the podocyte in the cytoplasmic aspect of the foot process membrane, adjacent to the insertion of the slit diaphragm (3-5). It links some of the slit diaphragm proteins through its PDZ (PSD-95/discs-large/ZO-1) domains to the actin cytoskeleton. Thus, changes in the expression or properties of ZO-1 may accompany renal diseases associated with proteinuria, including diabetic nephropathy. Studies in humans and experimental animals highlighted the importance of poor glycemic control in the development of the functional and structural changes in the kidney during the evolution of diabetic nephropathy (1,6 -10). Hyperglycemia and exposure of cultured cells to high glucose induce phenotypic modifications of cells that result in tissue injury (11)(12)(13)(14)(15). This study explored the expression of ZO-1 in two models of diabetes and the effect of glucose on ZO-1 expression and phosphorylation in cultured rat glomerular epithelial cells (GECs). RESEARCH DESIGN AND METHODSAnimals were used in accordance with ...

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Section: Zo-1 and Diabetic Nephropathymentioning

confidence: 99%

ZΟ-1 Expression and Phosphorylation in Diabetic Nephropathy

et al. 2006

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“…Ohtake et al reported that application of PLA to nasal mucosa changed the location of tight junction-related proteins, such as occludin and ZO-1, from the plasma membrane to the cytoplasm. 26) Another polycations, protamine, also showed that the expression and distribution of claudin-1 and occludin were altered in MDCK monolayers. 27) Thus, tight junction-associated proteins in ocular surface tissues, such as the cornea and conjunctiva, are likely to be disassembled in the presence of polycations like PLA.…”

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confidence: 99%

Effects of Poly-L-arginine on the Permeation of Hydrophilic Compounds through Surface Ocular Tissues

Nemoto¹,

Takahashi²,

Kobayashi³

et al. 2006

Biological & Pharmaceutical Bulletin

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Drugs that are topically applied to the cul-de-sac enter the intraocular area via both corneal and noncorneal pathways which are composed of cornea and conjunctiva-sclera, respectively.1) Although the corneal pathway is thought to be the primary route of intraocular entry for most drugs, penetration via the noncorneal pathway can also be important for the penetration of drugs which are poorly absorbed across the cornea. 2,3) In addition, the noncorneal pathway could be more important than the corneal pathway for the delivery of drugs to the posterior segment of the eye. 2,3)The first barrier to intraocular entry via the corneal and noncorneal pathways is the cornea and conjunctiva, respectively. These epithelial tissues are known to have tight junctions that act as a barrier in the paracellular spaces. 4,5) Because of this barrier structure in the epithelia, insufficient drug may be absorbed after instillation. Thus, subconjunctival and intravitreous injections are generally used in ocular pharmacotherapy.1) These invasive methods may not be acceptable to many patients and could potentially increase the risk of infection.6) In order to obtain a simpler and more acceptable form of application, the development of effective instilled formulations would be a major improvement. A number of ocular penetration enhancers, including calcium chelator (EDTA) and bile salts, have already been investigated, and it was found that those enhancers increased the apparent permeability coefficient (Papp) of FITC-dextran (MW: 4000, FD-4) by 2.9-to 15.5-fold in excised cornea and conjunctiva.7) However, these enhancers might not be acceptable from a safety point of view, because the following were observed: severe epithelial damage, such as a change in cellular morphology produced by EDTA and deoxycholate in the cornea, 8) leakage of cytosolic proteins produced by EDTA in the rectum, 9) and permeabilization caused by dissolution of the plasma membrane produced by bile salts. 10)Recently, a novel type of penetration enhancer has been developed for mucosal drug delivery. Illum et al. 11) showed that the polycationic compound, chitosan, greatly enhanced the absorption of insulin across the nasal mucosa in rats and sheep. Basic amino acid polymers such as poly-L-arginine (PLA) and poly-L-lysine (PLL) are also shown to effectively increase permeability of various compounds in nasal, tracheal, and kidney epithelia. [12][13][14] Although it is reported that PLL induces potential epithelial damages in tracheal epithelium, 13) PLA appears to enhance the absorption of hydrophilic molecules in mucosal tissues without producing any obvious epithelial changes, such as morphological alteration, leakage of cytosolic proteins, and release of phospholipids from the plasma membrane.15,16) Thus, PLA is considered to be an ideal permeation enhancer for epithelial drug delivery including ocular delivery.The purpose of the present study was to examine the enhancing effect of PLA on the permeability of hydrophilic compounds through the ocular epithelia....

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“…administration of rhGH was also evaluated. Although the rhGH absorption across the nasal mucosa was almost the same between the 0.25 and 0.5 % (w/v) of PLA (15) and PLA (150), it increased dramatically when the PLA concentration was 1.0 % (w/v). This is likely because the rhGH did not dissolve completely in the dosage solution when the PLA concentration was 0.25 or 0.5 % (w/v), but it almost completely dissolved in the 1.0 % (w/v) PLA.…”

Section: Discussionmentioning

confidence: 93%

“…The rhGH solution was mixed with or without a PLA solution (0.5, 1.0, and 2.0 % (w/v), PLA (15) and PLA (150)) prepared with succinate buffer (50 mM, pH 4.4) at a ratio of 1 : 1, then the mixtures containing rhGH (2.91 mg/mL) and PLA (0.25, 0.5, and 1.0 % (w/v)) were prepared for administration.…”

Section: Formulation For Intranasal Delivery Saizenmentioning

confidence: 99%

“…11,13) In the studies on the mechanism of enhanced absorption by PLA in the mucosa, it has been found that PLA induces the enhanced paracellular permeability by the dispersion of tight junction proteins (claudin-4, occludin, zonula occludens-1 (ZO-1) and tricellulin) into the cytoplasm through the cell-cell junction. [14][15][16][17] In particular, it was known that occludin was important for macromolecular permeation among tight junction proteins. 18,19) In addition, it has been also demonstrated that adherens junction protein (E-cadherin, β-catenin) is dispersed by PLA from the cell-cell junction.…”

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Development of a Transnasal Delivery System for Recombinant Human Growth Hormone (rhGH): Effects of the Concentration and Molecular Weight of Poly-L-arginine on the Nasal Absorption of rhGH in Rats

Kawashima

Uchida

Yamaki

et al. 2016

Biological & Pharmaceutical Bulletin

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A novel system for delivering recombinant human growth hormone (rhGH) that is noninvasive and has a simple method of administration is strongly desired to improve the compliance of children. The aim of this study was to investigate the potential for the intranasal (i.n.) co-administration of rhGH with poly-L-arginine (PLA) as a novel delivery system by evaluating the effects of the concentration and molecular weight of PLA on the nasal absorption of rhGH. The influence of the formation of insoluble aggregates and a soluble complex in the dosage formulation on nasal rhGH absorption was also evaluated by size-exclusion chromatography and ultrafiltration. PLA enhanced the nasal absorption of rhGH at each concentration and molecular weight examined. Nasal rhGH absorption increased dramatically when the PLA concentration was 1.0 % (w/v) due to the improved solubility of rhGH in the formulation. A delay in rhGH absorption was observed when the molecular weight of PLA was increased. This appeared to be because the increase in molecular weight caused the formation of a soluble complex. It seems that the PLA concentration affects the absorptionenhancing effect on rhGH, while the molecular weight of PLA affects the time when the maximum plasma rhGH concentration was reached (T max ) of rhGH after i.n. administration, mainly because of the interactions among rhGH, PLA, and additives. Therefore, the transnasal rhGH delivery system using PLA is considered to be a promising alternative to subcutaneous (s.c.) injection if these interactions are sufficiently controlled.Key words poly-L-arginine; nasal absorption; enhancer; recombinant human growth hormone Most peptide and protein medicinal drugs are administered clinically by injection. This invasive administration with a needle and syringe is associated with various problems, such as pain for the patients, the risk of trouble at the injection site, increased medical waste and the potential for complicated operation. Hence, an alternative route of administration is strongly desired. There have been numerous studies on the transmucosal delivery of peptide and protein drugs, such as oral, nasal, and pulmonary administration.1-3) However, these types of drugs are rapidly destroyed by the enzymes present in various organs, especially in the gastrointestinal tract, and have a limited permeation across the mucous membrane. Therefore, these drugs have low bioavailability.Recombinant human growth hormone (rhGH), a single polypeptide chain of 191 amino acids and a molecular mass of 22.1 kDa, is used to treat short stature in children caused by growth hormone deficiency, Turner's syndrome, or chronic renal failure. At present, rhGH is administered daily by subcutaneous (s.c.) injections. Therefore, compliance with rhGH therapy is often low, because children hate the pain caused by such injections. The compliance of patients affects the therapeutic outcome, because it is necessary to regularly administer injections of rhGH for the limited period of growth. 4,5) Hence, a novel rhGH deliver...

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Poly-l-Arginine Enhances Paracellular Permeability via Serine/Threonine Phosphorylation of ZO-1 and Tyrosine Dephosphorylation of Occludin in Rabbit Nasal Epithelium

Abstract: We conclude that poly-L-Arg enhances the paracellular permeability of FD-4 (i.e., macromolecules), at least, by way of both serine/threonine phosphorylation of ZO-1 and tyrosine dephosphorylation of occludin in rabbit nasal epithelium.

Cited by 62 publications

References 33 publications

ZΟ-1 Expression and Phosphorylation in Diabetic Nephropathy

ZΟ-1 Expression and Phosphorylation in Diabetic Nephropathy

Effects of Poly-L-arginine on the Permeation of Hydrophilic Compounds through Surface Ocular Tissues

Development of a Transnasal Delivery System for Recombinant Human Growth Hormone (rhGH): Effects of the Concentration and Molecular Weight of Poly-L-arginine on the Nasal Absorption of rhGH in Rats

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