Novel Biocompatible Adhesive for Intrarenal Embedding and Endoscopic Removal of Small Residual Fragments after Minimally Invasive Stone Treatment in an Ex Vivo Porcine Kidney Model: Initial Evaluation of a Prototype.

PURPOSE: Residual fragments related to endoscopic intracorporeal lithotripsy are a challenging problem. The impact of residual fragments remains a subject of discussion and growing evidence highlights that they have a central role in recurrent stone formation. Therefore, we developed a novel bioadhesive system for intrarenal embedding and retrieval of residual fragments after endoscopic lithotripsy in an ex vivo porcine kidney model. MATERIALS AND METHODS: In a standardized setting 30 human stone fragments 1 mm or less were inserted in the lower pole of an ex vivo porcine kidney model. We assessed the extraction efficacy of flexible ureteroscopy using the bioadhesive system in 15 preparations and a conventional retrieval basket in 15. Outcomes were compared regarding the endoscopic and macroscopic stone-free rate, and overall time of retrieval. RESULTS: Embedding and retrieving the residual fragment-bioadhesive complex were feasible in all trial runs. We observed no adverse effects such as adhesions between the adhesive and the renal collecting system or the instruments used. The stone-free rate was 100% and 60% in the bioadhesive and conventional retrieval groups, respectively (p = 0.017). Mean retrieval time was significantly shorter at 10 minutes 33 seconds vs 36 minutes 56 seconds in the bioadhesive group vs the conventional group (p = 0.001). CONCLUSIONS: This novel method involving adhesive based complete removal of residual fragments from the collecting system has proved to be feasible. Our evaluation in a porcine kidney model revealed that this technology performed well. Further tests, including inpatient studies, are required to thoroughly evaluate the benefit and potential drawbacks of bioadhesive based extraction of residual fragments after intracorporeal lithotripsy.

Glutathione-Dependent Detoxification Processes in Astrocytes.

Astrocytes have a pivotal role in brain as partners of neurons in homeostatic and metabolic processes. Astrocytes also protect other types of brain cells against the toxicity of reactive oxygen species and are considered as first line of defence against the toxic potential of xenobiotics. A key component in many of the astrocytic detoxification processes is the tripeptide glutathione (GSH) which serves as electron donor in the GSH peroxidase-catalyzed reduction of peroxides. In addition, GSH is substrate in the detoxification of xenobiotics and endogenous compounds by GSH-S-transferases which generate GSH conjugates that are efficiently exported from the cells by multidrug resistance proteins. Moreover, GSH reacts with the reactive endogenous carbonyls methylglyoxal and formaldehyde to intermediates which are substrates of detoxifying enzymes. In this article we will review the current knowledge on the GSH metabolism of astrocytes with a special emphasis on GSH-dependent detoxification processes.

Arsenite stimulates glutathione export and glycolytic flux in viable primary rat brain astrocytes.

Intoxication with inorganic arsenicals leads to neuropathies and impaired cognitive functions. However, little is known so far on the cellular targets that are involved in the adverse effects of arsenite to brain cells. To test whether arsenite may affect neural glucose and glutathione (GSH) metabolism, primary astrocyte cultures from rat brain were used as a model system. Exposure of cultured astrocytes to arsenite in concentrations of up to 0.3mM did not compromise cell viability during incubations for up to 6h, while 1mM arsenite damaged the cells already within 2h after application. Determination of cellular arsenic contents of astrocytes that had been incubated for 2h with arsenite revealed an almost linear concentration-dependent increase in the specific cellular arsenic content. Exposure of astrocytes to arsenite stimulated the export of GSH and accelerated the cellular glucose consumption and lactate production in a time- and concentration-dependent manner. Half-maximal stimulation of GSH export and glycolytic flux were observed for arsenite in concentrations of 0.1mM and 0.3mM, respectively. The arsenite-induced stimulation of both processes was abolished upon removal of extracellular arsenite. The strong stimulation of GSH export by arsenite was prevented by MK571, an inhibitor of the multidrug resistance protein 1, suggesting that this transporter mediates the accelerated GSH export. In addition, presence of MK571 significantly increased the specific cellular arsenic content, suggesting that Mrp1 may also be involved in arsenic export from astrocytes. The data observed suggest that alterations in glucose and GSH metabolism may contribute to the reported adverse neural consequences of intoxication with arsenite.

Antiretroviral protease inhibitors accelerate glutathione export from viable cultured rat neurons.

Antiretroviral protease inhibitors are crucial components of the antiretroviral combination therapy that is successfully used for the treatment of patients with HIV infection. To test whether such protease inhibitors affect the glutathione (GSH) metabolism of neurons, cultured cerebellar granule neurons were exposed to indinavir, nelfinavir, lopinavir or ritonavir. In low micromolar concentrations these antiretroviral protease inhibitors did not acutely compromise the cell viability, but caused a time- and concentration-dependent increase in the accumulation of extracellular GSH which was accompanied by a matching loss in cellular GSH. The stimulating effect by indinavir, lopinavir and ritonavir on GSH export was immediately terminated upon removal of the protease inhibitors, while the nelfinavir-induced stimulated GSH export persisted after washing the cells. The stimulation of neuronal GSH export by protease inhibitors was completely prevented by MK571, an inhibitor of the multidrug resistance protein 1, suggesting that this transporter mediates the accelerated GSH export during exposure of neurons to protease inhibitors. These data suggest that alterations in brain GSH metabolism should be considered as potential side-effects of a treatment with antiretroviral protease inhibitors.

8-Hydroxy-efavirenz, the primary metabolite of the antiretroviral drug Efavirenz, stimulates the glycolytic flux in cultured rat astrocytes.

In active antiretroviral therapy antiretroviral drugs are employed for the restoration of a functional immune system in patients suffering from the acquired immunodeficiency syndrome. However, potential adverse effects of such compounds to brain cells are discussed in connection with the development of neurocognitive impairments in patients. To investigate potential effects of antiretroviral drugs on cell viability and the glycolytic flux of brain cells, astrocyte-rich primary cultures were exposed to various antiretroviral compounds, including the non-nucleoside reverse transcriptase inhibitor efavirenz. In a concentration of 10 µM, neither efavirenz nor any of the other investigated antiretroviral compounds acutely compromised the cell viability nor altered glucose consumption or lactate production. In contrast, the primary metabolite of efavirenz, 8-hydroxy-efavirenz, stimulated the glycolytic flux in viable astrocytes in a time- and concentration-dependent manner with half-maximal and maximal effects at concentrations of 5 and 10 µM, respectively. The stimulation of glycolytic flux by 8-hydroxy-efavirenz was not additive to that obtained for astrocytes that were treated with the respiratory chain inhibitor rotenone and was abolished by removal of extracellular 8-hydroxy-efavirenz. In a concentration of 10 µM, 8-hydroxy-efavirenz and efavirenz did not affect mitochondrial respiration, while both compounds lowered in a concentration of 60 µM significantly the oxygen consumption by mitochondria that had been isolated form cultured astrocytes, suggesting that the stimulation of glycolytic flux by 8-hydroxy-efavrienz is not caused by direct inhibition of respiration. The observed alteration of astrocytic glucose metabolism by 8-hydroxy-efavirenz could contribute to the adverse neurological side effects reported for patients that are chronically treated with efavirenz-containing medications.

The antiretroviral protease inhibitor ritonavir accelerates glutathione export from cultured primary astrocytes.

Antiretroviral protease inhibitors are a class of important drugs that are used for the treatment of human immunodeficiency virus infections. Among those compounds, ritonavir is applied frequently in combination with other antiretroviral protease inhibitors, as it has been reported to boost their therapeutic efficiency. To test whether ritonavir affects the viability and the glutathione (GSH) metabolism of brain cells, we have exposed primary astrocyte cultures to this protease inhibitor. Application of ritonavir in low micromolar concentrations did not compromise cell viability, but caused a time- and concentration-dependent loss of GSH from the cells which was accompanied by a matching increase in the extracellular GSH content. Half-maximal effects were observed for ritonavir in a concentration of 3 µM. The ritonavir-induced stimulated GSH export from astrocytes was completely prevented by MK571, an inhibitor of the multidrug resistance protein 1. In addition, continuous presence of ritonavir was essential to maintain the stimulated GSH export, since removal of ritonavir terminated the stimulated GSH export. Ritonavir was more potent to stimulate GSH export from astrocytes than the antiretroviral protease inhibitors indinavir and nelfinavir, but combinations of ritonavir with indinavir or nelfinavir did not further stimulate astrocytic GSH export compared to a treatment with ritonavir alone. The strong effects of ritonavir and other antiretroviral protease inhibitors on the GSH metabolism of astrocytes suggest that a chronic treatment of patients with such compounds may affect their brain GSH metabolism.

The antiretroviral protease inhibitors indinavir and nelfinavir stimulate Mrp1-mediated GSH export from cultured brain astrocytes.

Combinations of antiretroviral drugs are successfully used for the treatment of acquired immune deficiency syndrome and reduce the incidence of severe human immunodeficiency virus (HIV)-associated dementia. To test whether such drugs affect the GSH metabolism of brain cells, we have exposed astrocyte-rich primary cultures to various antiretroviral compounds. Treatment of the cultures with the protease inhibitors indinavir or nelfinavir in low micromolar concentrations resulted in a time- and concentration-dependent depletion of cellular GSH from viable cells which was accompanied by a matching increase in the extracellular GSH content. In contrast, the reverse transcriptase inhibitors zidovudine, lamivudine, efavirenz or nevirapine did not alter cellular or extracellular GSH levels. Removal of indinavir from the medium by washing the cells terminated the stimulated GSH export immediately, while the nelfinavir-induced accelerated GSH export was maintained even after removal of nelfinavir. The stimulation of the GSH export from viable astrocytes by indinavir or nelfinavir was completely prevented by the application of MK571, an inhibitor of the multidrug resistance protein 1. These data demonstrate that indinavir and nelfinavir stimulate multidrug resistance protein 1-mediated GSH export from viable astrocytes and suggest that treatment of patients with such inhibitors may affect the GSH homeostasis in brain.