Distribution and characterization of the opioid octapeptide met5-enkephalin-arg6-gly7-leu8 in the gastrointestinal tract of the rat

Summary The distribution and characterization of the opioid octapeptide met⁵-enkephalin-arg⁶-gly⁷-leu⁸ (met⁵-enk-arg⁶-gly⁷-leu⁸) within the gastrointestinal tract of the rat has been determined by immunohistochemistry and radioimmunoassay by use of a newly developed antibody to met⁵-enk-arg⁶-gly⁷-leu⁸. With both techniques, met⁵-enk-arg⁶-gly⁷-leu⁸-immunoreactivity (met⁵-enk-arg⁶-gly⁷-leu⁸IR) was detected in all regions of the gastrointestinal (GI) tract except the esophagus. The highest concentration of immunoreactive met⁵-enk-arg⁶-gly⁷-leu⁸ was observed in the colon, while intermediate concentrations were found in the stomach, duodenum, jejunum, and ileum. Immunostained somata were observed chiefly in the myenteric plexus; immunostained processes were present primarily in the myenteric plexus and the circular muscle layer. This distribution pattern is similar to that previously observed with antiserum to met⁵-enkephalin-arg⁶-phe⁷ (met⁵-enk-arg⁶phe⁷). Chromatographic analysis of met⁵-enk-arg⁶-gly⁷leu⁸-immunoreactive peptides extracted from the GI tract revealed the presence of an immunoreactive peptide of high molecular weight which accounted for approximately three-quarters of met⁵-enk-arg⁶-gly⁷-leu⁸-IR in both stomach and colon. These findings suggest a role for peptides related to the octapeptide met⁵-enk-arg⁶-gly⁷-leu⁸ in the regulation of GI function.     

Correlation between automated DNA ploidy measurements of Hürthle-cell tumors and their histopathologic and clinical features

The treatment of Hürthle-cell tumors of the thyroid is controversial because of their rarity and the inconsistent histopathologic criteria for their diagnosis. In order to obtain more objective criteria for the management of Hürthle-cell tumors, the nuclear DNA content of cells from 20 cases was measured with the MicroTICAS system and the correlation between the DNA distribution patterns and the clinical and histopathologic findings was evaluated. Three main DNA patterns were found: euploid, polyploid and aneuploid. The euploid or polyploid Hürthle-cell tumors came from patients who did not develop distant metastases or recurrence whereas the aneuploid variants came from patients who died of their disease and/or developed distant metastases and recurrence. Various correlation analyses were performed between DNA ploidy and age, sex, size of tumor, growth pattern, pleomorphism, invasion and metastases. Our data suggests that an aneuploid DNA pattern or one with a large percentage of aneuploid nuclei with DNA content exceeding 5N may predict eventual metastases or recurrence from Hürthle-cell tumor.     

Refined 2 A X-ray crystal structure of porcine pancreatic kallikrein A, a specific trypsin-like serine proteinase. Crystallization, structure determination, crystallographic refinement, structure and its comparison with bovine trypsin

Porcine pancreas kallikrein A has been crystallized in the presence of the small inhibitor benzamidine, yielding tetragonal crystals of space group P4₁2₁2 containing two molecules per asymmetric unit. X-ray data up to 2·05 Å resolution have been collected using normal rotation anode as well as synchrotron radiation. The crystal structure of benzamidine-kallikrein has been determined using multiple isomorphous replacement techniques, and has subsequently been refined to a crystallographic R-value of 0·220 by applying a diagonal matrix least-squares energy constraint refinement procedure.Both crystallographically independent kallikrein molecules 1 and 2 are related by a non-integral screw axis and form open, heterologous “dimer” structures. The root-mean-square deviation of both molecules is 0·37 Å for all main-chain atoms. This value is above the estimated mean positional error of about 0·2 Å and reflects some significant conformational differences, especially at surface loops. The binding site of molecule 1 in the asymmetric unit is in contact with residues of molecule 2, whereas the binding site of the latter is free and accessible to the solvent. In both molecules the characteristic “kallikrein loop”, where the peptide chain of kallikrein A is cleaved, is only partially traceable. The carbohydrate attached to Asn95 in this loop, although detectable chemically, is not defined.A comparison of the refined structures of porcine kallikrein and bovine trypsin indicates spatial homology for these enzymes. The root-mean-square difference is 0·68 Å if we compare only main-chain atoms of internal segments. Remarkably large deviations are found in some external loops most of which surround the binding site and form a more compact rampart around it in kallikrein than in trypsin. This feature might explain the strongly reduced activity and accessibility of kallikrein towards large protein substrates and inhibitors (e.g. as shown by the model-building experiments on inhibitor complexes reported by Chen &; Bode. 1983).The conformation of the active site residues is very similar in both enzymes. Tyr99 of kallikrein, which is a leucyl residue in trypsin, protrudes into the binding site and interferes with the binding of peptide substrates (Chen &; Bode. 1983). The kallikrein specificity pocket is significantly enlarged compared with trypsin due to a longer peptide segment, 217 to 220, and to the unique outwards orientation of the carbonyl group of cis-Pro219. Further, the side-chain of Ser226 in porcine kallikrein, which is a glycyl residue in trypsin, partially covers Asp 189 at the bottom of the pocket. These features considerably affect the binding geometry and strength of binding of benzamidine.     

High-resolution color video cytophotometry

Comparison was made between cytophotometric measurements obtained using two data acquisition systems, one a microphotometer and the other a rapid video camera system, to ascertain whether the degradation of data with the faster video acquisition system still results in recorded images of sufficient quality to permit computer discrimination between cells of very similar appearance. Normal-appearing intermediate cells from cases with normal cytology and those from patients with dysplasia or malignant disease, as well as the subvisual markers within these cells that have rendered them capable of cytophotometric discrimination, were used for the study. Comparison of the data recorded by the two systems indicates that the diagnostic information is preserved in the change-over to a full-field, video-rate scanning system, with differences in the data caused primarily by differences in the spectral response of the two systems. This was reflected in the substantial differences observed in the color-related features and the lesser differences seen in the textural features, while the morphometric features (outline and shape) were virtually unaffected. The differences were primarily expressed on a cell-to-cell basis; in sets of about 300 cells, which would be used in patient-to-patient comparisons, the feature values showed remarkable consistency between the two systems.     

Enzymbestimmungen in Erythrocyten bei Kindern mit Down-Syndrom

Zusammenfassung Bei 20 Kleinkindern mit gesicherter freier Trisomie 21 wurden die Aktivitäten von 21 Enzymen des Energie- und des Glutathionstoffwechsels in isolierten Erythrocyten gemessen. Die Aktivitäten der Fructose-6-phosphatkinase, der Lactatdehydrgenase, der Glucose-6-phosphatdehydrogenase, der Glutathionreduktase (NADP) und der Glutamat-Oxalacetat-Transaminase wurden gegenüber gleichaltrigen Kontrollen signifikant erhöht gefunden. Lediglich die etwa 50%ige Steigerung der Fructose-6-phosphatkinase-Aktivität kann im Sinne eines Gen-Dosis-Effektes gedeutet werden.

---

Enzyme activities in the erythrocytes of children with Down's syndrom

Summary The activities of 21 enzymes of the energy supplying and the glutathione metabolism were estimated in isolated erythrocytes from children with Down's syndrome aged from 1 to 5 1/2 years. All 20 patients were trisomic for chromosome 21. The following enzymes were estimated by means of optic assays: All glycolytic enzymes, two enzymes of the hexosephosphate shunt and two of the citric cycle, NADP- and NAD-dependent glutathione reductase, glutamic-oxaloacetic transaminase, adenylate kinase, and Mg²⁺-activated ATPase.The activities of five enzymes, i.e. phosphofructokinase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase, glutathione reductase (NADP), and glutamic-oxaloacetic transaminase, were significantly elevated as compared to age-matched controls. The 50% increase of phosphofructokinase-activity is assumed to provide suggestive evidence for a controlling gene-locus on chromosome 21. The remaining findings are not considered to be directly related to the chromosomal abnormality.

---

---

Direcktor: Prof. Dr. H.-R. Wiedemann

---

Nach einem Vortrag auf der 10. Tagung der Gesellschaft für Anthropologie und Human-genetik, Königstein (Ts.), 22.–25. 10. 1967.

---

Die Untersuchungen wurden durch die dankenswerte Unterstützung der Deutschen Forschungsgemeinschaft ermöglicht.     

Image understanding system for histopathology

An image understanding machine vision system for histological diagnoses is based on three interacting expert systems: a diagnostic expert system utilizing terms familiar to pathologists, an interpretive expert system relating human diagnostic concepts to computable histometric features and a scene segmentation expert system which extracts the diagnostic information from the imagery. The control software for the image understanding system resides on a multiprocessor computer. This article details measures to maintain system efficiency and to accommodate the requirements of interprocess communication and processing task scheduling.     

Crystals of wild-type, mutated, derivatized and complexed 50 S ribosomal subunits from Bacillus stearothermophilus suitable for X-ray analysis

Three-dimensional single crystals of wild-type and mutated 50 S ribosomal subunits from Bacillus stearothermophilus, as well as crystals of reconstituted subunits containing heavy-atom clusters and complexes of these subunits with tRNA and a short nascent polypeptide chain, were grown from polyethylene glycol in the presence of salts at low concentrations. Within experimental error, all these crystals are isomorphous, packed with monoclinic symmetry (C2) in unit cells of a = 300 A, b = 546 A, c = 377 (+/- 1%) A and beta = 112 degrees. Using synchrotron radiation at 85 to 100 K they diffract to 11 A resolution and can be irradiated for hours without disintegrating, so that a complete data set could be collected from a single crystal.