Expression of human cationic trypsinogen (PRSS1) in murine acinar cells promotes pancreatitis and apoptotic cell death.

Hereditary pancreatitis (HP) is an autosomal dominant disease that displays the features of both acute and chronic pancreatitis. Mutations in human cationic trypsinogen (PRSS1) are associated with HP and have provided some insight into the pathogenesis of pancreatitis, but mechanisms responsible for the initiation of pancreatitis have not been elucidated and the role of apoptosis and necrosis has been much debated. However, it has been generally accepted that trypsinogen, prematurely activated within the pancreatic acinar cell, has a major role in the initiation process. Functional studies of HP have been limited by the absence of an experimental system that authentically mimics disease development. We therefore developed a novel transgenic murine model system using wild-type (WT) human PRSS1 or two HP-associated mutants (R122H and N29I) to determine whether expression of human cationic trypsinogen in murine acinar cells promotes pancreatitis. The rat elastase promoter was used to target transgene expression to pancreatic acinar cells in three transgenic strains that were generated: Tg(Ela-PRSS1)NV, Tg(Ela-PRSS1*R122H)NV and Tg(Ela-PRSS1*N29I)NV. Mice were analysed histologically, immunohistochemically and biochemically. We found that transgene expression is restricted to pancreatic acinar cells and transgenic PRSS1 proteins are targeted to the pancreatic secretory pathway. Animals from all transgenic strains developed pancreatitis characterised by acinar cell vacuolisation, inflammatory infiltrates and fibrosis. Transgenic animals also developed more severe pancreatitis upon treatment with low-dose cerulein than controls, displaying significantly higher scores for oedema, inflammation and overall histopathology. Expression of PRSS1, WT or mutant, in acinar cells increased apoptosis in pancreatic tissues and isolated acinar cells. Moreover, studies of isolated acinar cells demonstrated that transgene expression promotes apoptosis rather than necrosis. We therefore conclude that expression of WT or mutant human PRSS1 in murine acinar cells induces apoptosis and is sufficient to promote spontaneous pancreatitis, which is enhanced in response to cellular insult.

Free radicals and the pancreatic acinar cells: role in physiology and pathology.

Reactive oxygen and nitrogen species (ROS and RNS) play an important role in signal transduction and cell injury processes. Nitric oxide synthase (NOS)-the key enzyme producing nitric oxide (NO)-is found in neuronal structures, vascular endothelium and, possibly, in acinar and ductal epithelial cells in the pancreas. NO is known to regulate cell homeostasis, and its effects on the acinar cells are reviewed here. ROS are implicated in the early events within the acinar cells, leading to the development of acute pancreatitis. The available data on ROS/RNS involvement in the apoptotic and necrotic death of pancreatic acinar cells will be discussed.

Rat hippocampal neurons maintain their own GABAergic synaptic transmission in culture.

The whole-cell patch-clamp technique was used to record monosynaptic inhibitory postsynaptic currents (IPSCs) from pairs of hippocampal neurons cultured for 2-3 weeks. The application of fresh physiological solution for 2-3 min reversibly reduced the amplitude of evoked GABAergic IPSCs to 72.5% of control value. The amplitude and frequency of spontaneous IPSCs decreased too. The depression of evoked IPSCs was significantly smaller or absent if conditioned solution was applied (physiological solution which had been previously in contact with neurons for 30 min). Currents evoked by exogenously applied GABA were unaffected by fresh solution. These results suggest that hippocampal neurons release some endogenous substance(s), by which they up regulate presynaptically their own inhibitory synaptic transmission.

Nifedipine-induced morphological differentiation of rat pheochromocytoma cells.

Extension of neuronal processes is fundamental for the establishment of the intricate network of the nervous system. The rat PC12 pheochromocytoma cell line is a well-known model system for the study of neuronal differentiation. In the present study the effect of a high-voltage-activated calcium channel blocker, nifedipine, on the cell differentiation was investigated. In cells cultured in a modified medium containing nifedipine (at 2.5 microM or 5.0 microM) a decrease of mitotic activity took place on the third day of culturing. An increase of total length and number of processes occurred on the fifth day. A higher concentration of nifedipine (10.0 microM) considerably suppressed vital functions of the cells. Retraction of processes together with blocking of proliferation activity was observed. In parallel with these changes, a reduction of [Ca2+]i from 100 +/- 6 nM to 50 +/- 2 nM was detected on the second day of cultivation in 2.5 microM and 5.0 microM nifedipine containing medium. At 10.0 microM concentration of nifedipine in the medium a decrease in [Ca2+]i was observed on the first day only (to 76 +/- 8 nM); then the level of free calcium concentration rose dramatically. The data obtained allow us to suggest that the decrease of [Ca2+]i during first two days of PC12 cells culturing in a nifedipine-containing medium could be a factor which stimulated their morphological differentiation.