Induction of anti-carbonic-anhydrase-II antibody causes renal tubular acidosis in a mouse model of Sjogren's syndrome.

BACKGROUND/AIM: We recently reported that renal tubular acidosis (RTA) in Sjogren's syndrome (SjS) is associated with high titers of an autoantibody against carbonic anhydrase (CA) II, an important enzyme in renal acid-base regulation. The purpose of this study was to determine whether a CA-II antibody could cause RTA in a mouse model of SjS. METHODS: PL/J mice were immunized with human CA II to induce CA II antibody formation, whereas controls were injected with phosphate-buffered saline and adjuvant. After 6 weeks, anti-CA-II antibody titers were measured, then ammonium chloride was administered orally for 1 week to detect any acidification defect. RESULTS: CA-II-immunized mice showed higher anti-CA-II antibody titers than control mice. Pathologically, lymphocytic and plasma cell infiltration was seen in the salivary glands and kidneys of CA-II-immunized mice, but not in controls. On acid loading, blood pH and urine pH decreased in both groups of mice, but the slope of urine pH versus blood pH was less steep in the CA-II-immunized mice, suggesting that these mice had an impaired ability to reduce their urine pH in the face of metabolic acidosis. CONCLUSION: CA-II-immunized mice had a urinary acidification defect, which may be similar to that seen in patients with SjS.

Effects of a sperminated gelatin on the nasal absorption of insulin.

The effects of a sperminated gelatin (SG), which was prepared as a candidate absorption enhancer by the addition of spermine to gelatin, on the nasal absorption of insulin, were examined in rats. The AUC of immuno-reactive insulin levels in the plasma after nasal administration of insulin were increased 5.3-fold by addition of 0.2% SG, and the plasma glucose levels fell in a manner dependent on the insulin levels. In Calu-3 cell monolayer permeation experiments, SG showed significant enhancing effects on 5(6)-carboxyfluorescein (CF), FITC-dextran (MW 4400, FD4) and insulin. Evaluation of the tight junctions in the Calu-3 cell monolayers based on the Renkin molecular sieving function suggests that the pore occupancy/length ratio of the permeation pathways for water-soluble molecules in the tight junctions increases, while the equivalent cylindrical pore radius is not changed by SG treatment. SG may transform the true tight junctions, which act as a barrier for water-soluble molecules, into pathways for CF and FD4 to increase their number. SG is a good candidate for a safe absorption enhancer to produce a slight modification of the permeability of the paracellular pathway of mucosal membranes, while retaining the sieving property of the epithelial membranes.

Effect of cationized gelatins on the paracellular transport of drugs through caco-2 cell monolayers.

Cationized gelatins, candidate absorption enhancers, were prepared by addition of ethylenediamine or spermine to gelatin and the effects of the resulting ethylenediaminated gelatin (EG) and sperminated gelatin (SG) on the paracellular transport of 5(6)-carboxyfluorescein (CF), FITC-dextran-4 (FD4), and insulin through caco-2 cell monolayers were examined. The Renkin function was used for characterization of the paracellular pathway and changes in the pore radius (R) and pore occupancy/length ratio (epsilon/L) calculated from the apparent permeability coefficients (P(app)) of CF and FD4 are discussed. Ethylenediaminetetraacetic acid (EDTA) increased the R of the caco-2 cell monolayer and the P(app) of all compounds examined was markedly increased by the addition of EDTA. On the other hand, EG and SG did not increase R and their enhancing effects were not as strong as those of EDTA. The increase in epsilon/L could be the enhancing mechanism for the cationized gelatins. The number of pathways for water-soluble drugs, such as CF and FD4, in the caco-2 monolayers could be increased by the addition of the cationized gelatins. The ratios of the permeability coefficients of insulin (observed/calculated based on the Renkin function) suggest that insulin undergoes enzymatic degradation during transport which is not inhibited by enhancers.

Mechanisms underlying the maintenance of muscle hypercontractility in a model of postinfective gut dysfunction.

BACKGROUND & AIMS: Acute gastroenteritis is a strong risk factor for the development of irritable bowel syndrome (IBS). We have developed an animal model in which transient acute infection leads to persistent muscle hypercontractility. Here, we investigate the mechanisms underlying the maintenance of this hypercontractility in the postinfective (PI) state. METHODS: Muscle contraction and messenger RNA (mRNA) or protein expression of cytokines were examined from jejunal longitudinal muscle cells of NIH Swiss mice infected with Trichinella spiralis or incubated with or without cytokines. RESULTS: During acute infection, interleukin (IL)-4 or IL-13, transforming growth factor (TGF)-beta1, and cyclooxygenase (COX)-2 were increased in the muscle layer ( P < .05). In the PI phase of the model, T helper (Th)2 cytokines returned to normal, but TGF-beta1 remained in the muscle ( P < .05). Exposure of muscle cells to IL-4 or IL-13 increased TGF-beta1 ( P < .01), COX-2 protein, and prostaglandin (PG)E 2 . Exposure of muscle cells to TGF-beta1 increased PGE 2 ( P < .05) and COX-2 protein. Incubation of tissue with IL-4, IL-13, TGF-beta1, or PGE 2 enhanced carbachol-induced muscle cell contractility ( P < .05). COX-2 inhibitor attenuated TGF-beta1-induced muscle hypercontractility ( P < .05). CONCLUSIONS: These results support the hypothesis that Th2 cytokines induce muscle hypercontractility during infection by a direct action on smooth muscle. The maintenance of hypercontractility results from Th2 cytokine-induced expression of TGF-beta1 and the subsequent up-regulation of COX-2 and PGE 2 at the level of the smooth muscle cell. We propose that PI gut dysfunction reflects mediator production in the neuromuscular tissues and that this may occur in PI-IBS.

Effect of aminated gelatin on the nasal absorption of insulin in rats.

Absorption enhancers, which increase the permeability of drugs through epithelial membranes without damaging them, are especially useful for intranasal administration of peptide drugs. In this study, aminated gelatins, candidate enhancers, having different numbers of amino groups were prepared from gelatin (H-gelatin, isoelectric point = 9.0, MW 100 kDa) and a partial gelatin hydrolysate (L-gelatin, isoelectric point = 8.0, MW 5 kDa), and the enhancing effects on the nasal absorption of insulin, used as a model peptide drug, and 5(6)-carboxyfluorescein (CF), a paracellular marker, were examined in rats. The enhancing effect on insulin and CF depends on the MW and number of amino groups. A high correlation between the enhancing effects on insulin and CF was observed and this suggests that an increase in the paracellular permeability is the mechanism governing the nasal absorption-enhancement of aminated gelatins, at least as far as insulin and CF are concerned. The enhancing mechanism might be shared with other cationic polymers having absorption-enhancing effects.