Improving the barrier function of damaged cultured urothelium using chondroitin sulfate

Abstract

Aims: To determine whether glycosaminoglycan (GAG) replenishment is able to improve recovery of a deficient urothelial barrier, chondroitin sulfate (CS) instillations were tested using an in vitro model. Porcine urothelial cells (Ucells) were terminally differentiated in culture conditions to construct a urothelial layer with a functional barrier. This layer was damaged to compromise barrier function to simulate a key characteristic of bladder pain syndrome/interstitial cystitis. The functional effect of subsequent treatment with CS was evaluated.Methods: Primary porcine Ucells were isolated and cultured on inserts. Differentiation of cells was evaluated with immunohistochemical analysis for the presence of umbrella cells, tight junctions and CS. Transepithelial electrical resistance (TEER) measurements were performed to evaluate barrier function. Protamine was used to simulate mild urothelial damage. CS 0.2% (vol/vol), a GAG, was subsequently instilled in the treatment group. The recovery of barrier function was further evaluated with TEER measurements. The Student t test was used for the analysis of results.

Chondroitin sulfate GAG-replenishment therapy and its effect on restoring the barrier function of urothelium in an experimental model for BPS/IC.

“…When confluency was reached, the medium was supplied with 5% fetal calf serum (FCS) and 2mM calcium chloride (Ca 2+ Cl) to induce terminal differentiation. [8] Terminal differentiation was confirmed with TEER measurements and immunofluorescence analyses.…”

“…[1,2] The primary aim of GAG therapy is to improve barrier function and recovery of the chronically damaged urothelium. [8] Currently clinical research still outweighs the efforts done to investigate the physiological mechanisms behind GAG therapy.…”

“…This implies mostly a single exposure of the urothelium to a harmful substance. [8] However, the chronic aspect of BPS/IC is one of its main features. This in-vitro study was aimed to 1) 1) to evaluate the therapeutic efficacy of CS replenishment therapy in a chronically damaged urothelial model and 2) establish the effects of chronic damage on the functional barrier and recovery of the bladder urothelium in a controlled environment.…”

A Systematic Review of Therapeutic Approaches Used in Experimental Models of Interstitial Cystitis/Bladder Pain Syndrome

“…To evaluate the effect of CS on the barrier function after induction of urothelial damage, Rozzenberg et al used terminally differentiated porcine urothelial cells, which are morphologically and functionally comparable with the same types of cells in a normal human urothelium. Treatment with CS significantly accelerated the recovery of the barrier function 7 h after acute damage with PS [ 31 ]. Rooney et al showed that high-molecular-weight HA significantly decreased TNFα- and PS-induced IL8 and IL6 production, increased sulfated GAG production, and decreased trans-epithelial permeability without altering tight junction protein expression in the HTB4 urothelial cell line [ 32 ].…”

MRI as a Tool to Assess Interstitial Cystitis Associated Bladder and Brain Pathologies

“…The urothelial GAG layer has a crucial role in delivering a permeability barrier to prevent penetration of urinary toxins and pathogens into the bladder wall [ 24 , 26 ]. In multiple animal models [ 27 , 28 , 29 ] as well as in cell culture models [ 30 ], disruption of the GAG layer results in an induction of permeability that can be restored to normal impermeability by treatment with GAGs [ 31 , 32 , 33 ]. Most convincing is that digestion of chondroitin sulfate on the bladder surface with the specific enzyme chondroitinase ABC alone can induce the same degree of permeability as treatment with protamine sulfate [ 33 ].…”