Multifactorial genesis of pancreatitis in primary hyperparathyroidism: evidence for "protective" (PRSS2) and "destructive" (CTRC) genetic factors.

OBJECTIVE: A relationship between primary hyperparathyroidism (pHPT) and pancreatitis has long been debated and remains a rare epiphenomenon. In a cohort of patients with pHPT and pancreatitis mutations in the serine protease inhibitor Kazal type I (SPINK1) and cystic fibrosis transmembrane conductance regulator (CFTR) genes, that increase the risk for pancreatitis have already been detected. Among the identification of additional pancreatitis-associtated mutations in the Chymotrypsin C gene (CTRC) it became clear that also protective genetic variants exist in the anionic trypsinogen gene (PRSS2) that decrease susceptibility for pancreatitis. Our aim was to detect either protective or inducing genetic factors in a large cohort of pHPT patients. METHODS: Among 1,259 patients with pHPT, 57 patients were identified with pancreatitis (4.5%). DNA was available from 31 patients (16 acute pancreatitis/15 chronic pancreatitis). These individuals and 100 patients with pHPT without pancreatitis were analysed for CTRC (p.R254W and p.K247_R254del) and PRSS2 (p.G191R) mutations using melting curve analysis and DNA sequencing or PCR and gel electrophoresis (in case of p.K247_R254del CTRC). RESULTS: 2 of 31 patients with pHPT and pancreatitis carried the CTRC p.R254W missense mutation (6.5%), while all 100 pHPT controls without pancreatitis showed no CTRC mutation (P=0.055). No further SPINK1 p.N34S (n=4) mutations were detected but the probability of either CTRC or SPINK1 mutations in pHPT patients with pancreatitis is high (P<0.05). 1 patient was trans-heterozygous ( SPINK1: N34S/ CTRC p.R254W). CTRC p.K247_R254del was not detected in both groups. PRSS2 (p.G191R) mutation was present in 1 patient with pancreatitis (3.2%) and in 6 pHPT controls (6%) (P=1). CONCLUSION: This study underlines the relevance of a genetic background in pHPT related pancreatitis. However, it only indicates that the CTRC (p.R254W) mutation might also contribute to the panel of mutations ( SPINK1 and CFTR) that have been formerly reported to elevate pancreatitis susceptibility in pHPT. Besides it suggests that protective genetic variants, i. e., p.G191R PRSS2, may contribute to the low prevalence of pancreatitis in pHPT patients.

A novel A121T mutation in human cationic trypsinogen associated with hereditary pancreatitis: functional data indicating a loss-of-function mutation influencing the R122 trypsin cleavage site.

BACKGROUND: The understanding of genetic risk factors for chronic pancreatitis increased in the last decade with the discovery of mutations in the cationic trypsinogen gene (PRSS1). The first mutation was detected at the R122 autocleavage site of the protein (R122H) and subsequently two other mutations in this region, R122C and V123M, were described that resulted in a similar phenotype of hereditary pancreatitis. This study reports a novel A121T mutation within this region and characterises the resulting molecular properties at the autocleavage site. METHODS: Blood samples of a PRSS1 A121T carrier family were analysed for PRSS1 mutations using melting point curve analysis, restriction endonucleases and DNA sequencing. Conformation dependent properties of the mutated sequence were analysed by molecular modelling. The autodegradation kinetic of the mutated trypsin sequence was measured by a novel fluorescence resonance energy transfer (FRET) assay using designed 11 amino acid peptides from PRSS1 aa 118-aa 127 containing the trypsin cleavage site at aa 122 coupled to a Dabcyl/EDANS FRET system. The kinetic of tryptic peptide cleavage was measured in a fluorescence enzyme linked immunosorbent assay (ELISA) reader. RESULTS: DNA sequencing revealed a novel G to A transition at position 133279 of the published genomic sequence (#U66061 GenBank). The mutation results in an amino acid substitution of alanine by threonine at position 121 (A121T) of the cationic trypsinogen. Four additional mutation carriers could be identified among the relatives while only the first patient developed chronic pancreatitis. Molecular modelling of PRSS1 A121T revealed a change in the bond pattern between the R122 region and the calcium binding loop, whereas FRET assays showed an increased trypsin cleavage rate with a reaction kinetic elevated by more than 80%. CONCLUSION: The novel PRSS1 A121T mutation highlights the surface exposed region PRSS1 A121-R122-V123 as a hotspot for hereditary pancreatitis associated trypsinogen mutations. Molecular modelling and FRET assays provide evidence for an A121T mutation dependent increase in susceptibility to trypsin digestion at the R122 cleavage site suggesting an enhanced autodegradation and a loss-of-function at the autocleavage site.

[In vitro effects of diclofenac and selective cyclooxygenase-2 inhibitors on prostaglandin release from inflamed bursa subacromialis tissue in patients with subacromial syndrome]. / Effekte von Diclofenac und selektiven Cyclooxygenase-(COX-)2-Inhibitoren auf die Prostaglandinfreisetzung aus entzündetem Bursa-subacromialis-Gewebe von Patienten mit Subakromialsyndrom in vitro.

BACKGROUND: To compare the in vitro effects of selective COX-2 inhibitors (L-745,337, NS-398 and DFU) and of COX-unspecific diclofenac on release of PGE(2 )and 6-keto-PGF(1alpha) from inflamed bursa subacromialis tissue (IBST) obtained from a total of 35 patients with shoulder impingement syndrome (SIS). PATIENTS AND METHODS: Bursal specimens were incubated in the presence of drugs (0.01-1000 microM) for 20 min and 16 h. RESULTS: After 20 min 10 microM diclofenac significantly inhibited formation of PGE(2) and 6-keto-PGF(1alpha), whereas L-745,337 and NS-398 (10-1000 microM) induced significant inhibition only at concentrations > or =100 microM. In contrast to equimolar diclofenac, DFU (0.01-10 microM) induced no inhibition of bursal PGE(2) release but a dose-dependent, although statistically not significant inhibition after 16 h. The inhibitory potency of diclofenac (0.01-10 microM) was even more increased during long-term incubation showing greater inhibition than DFU at all concentrations studied. CONCLUSION: The data suggest that in IBST in SIS in vitro the majority of PG is generated via the COX-1 pathway.

Pinch-plasma radiation source for extreme-ultraviolet lithography with a kilohertz repetition frequency.

An extreme-ultraviolet radiation source based on a xenon pinch plasma is discussed with respect to the demands on a radiation source for extreme-ultraviolet lithography. Operation of the discharge in a self-igniting-plasma mode and omitting a switch permits a very effective and low-inductive coupling of the electrically stored energy to the electrode system. The xenon plasma exhibits broadband emission characteristics that offer radiation near 11 and 13 nm. Both wavelengths are useful in combination with beryllium- and silicon-based multilayer mirrors. The plasma emits approximately 74 mW/sr at 11.5 nm and 40 mW/sr at 13.5 nm in a bandwidth of 2% when operated at a repetition frequency of 120 Hz. The source size is less than 500 microm in diameter (FWHM) when viewed from the axial direction. The pulse-to-pulse stability is better than 3.6%. First results with a repetition rate of as much as 6 kHz promise the possibility of scaling to the required emission power for extreme-ultraviolet lithography.

Highly repetitive, extreme-ultraviolet radiation source based on a gas-discharge plasma.

An extreme-ultraviolet (EUV) radiation source near the 13-nm wavelength generated in a small (1.1 J) pinch plasma is presented. The ignition of the plasma occurs in a pseudosparklike electrode geometry, which allows for omitting a switch between the storage capacity and the electrode system and for low inductive coupling of the electrically stored energy to the plasma. Thus energies of only a few joules are sufficient to create current pulses in the range of several kiloamperes, which lead to a compression and a heating of the plasmas to electron densities of more than 10(17) cm(-3) and temperatures of several tens of electron volts, which is necessary for emission in the EUV range. As an example, the emission spectrum of an oxygen plasma in the 11-18-nm range is presented. Transitions of beryllium- and lithium-like oxygen ions can be identified. Current waveform and time-resolved measurements of the EUV emission are discussed. In initial experiments a repetitive operation at nearly 0.2 kHz could be demonstrated. Additionally, the broadband emission of a xenon plasma generated in a 2.2-J discharge is presented.

Laser-produced lithium plasma as a narrow-band extended ultraviolet radiation source for photoelectron spectroscopy.

Extended ultraviolet (EUV) emission characteristics of a laser-produced lithium plasma are determined with regard to the requirements of x-ray photoelectron spectroscopy. The main features of interest are spectral distribution, photon flux, bandwidth, source size, and emission duration. Laser-produced lithium plasmas are characterized as emitters of intense narrow-band EUV radiation. It can be estimated that the lithium Lyman-alpha line emission in combination with an ellipsoidal silicon/molybdenum multilayer mirror is a suitable EUV source for an x-ray photoelectron spectroscopy microscope with a 50-meV energy resolution and a 10-mum lateral resolution.

Soft-x-ray imaging with toroidally curved thallium Acid phthalate crystals in the water window.

Two-dimensional imaging in the wavelength region of the water window (N Ly alpha) is demonstrated with a toroidally curved thallium acid phthalate crystal. Direct imaging of a plasma pinch line, as well as imaging of a mesh with plasma backlighting in which the same source is used, is shown. For verification, the results are compared with the results by other methods. An absolute intensity calibration of the images is demonstrated with theoretical data for the reflection properties of the crystal. This advance in two-dimensional imaging was possible owing to the recent progress in the precise bending of acid phthalate crystals to defined concave shapes of high quality (sphere, toroid). With these crystals, the two-dimensional imaging method is pushed to the wavelength of 2.66 nm, which is well inside the water window.

Pinch plasma source for x-ray microscopy with nanosecond exposure time.

The strong demand for bright, compact, and inexpensive sources for x-ray microscopy has stimulated the development of flash x-ray sources. In this paper, the requirements for such a source are analyzed under boundary conditions given by the concept of an imaging x-ray microscope using mirror condenser and Fresnel zone plates for high-resolution imaging. It is found that the Lyman-α (1s-2p) line of hydrogen-like nitrogen (N VII) at λ = 2.48 nm emitted from a nonequilibrium plasma of about 200 eV temperature and 1020 cm-3 electron density is best suited. These conditions are achieved in medium-current pinch-plasma devices. Using detailed numerical simulation of the physical processes of such a device, optimization criteria for the integrated spectral brightness (ISB) are found. Measurements of the ISB confirm these optimization criteria. The results show that the spectral emission characteristics of an optimized pinch plasma souce are compatible with the demands of the mentioned x-ray microscopy concept. These emission characteristics are compared with laser-produced plasma sources. Using the optimized source with an ISB exceeding 0.6 µJ/(µm2 sr) in a 10-20 ns pulse, wet biological samples are imaged with about 0.1 µm lateral resolution.