Destabilization of lens protein conformation by glutathione mixed disulfide

Mixed disulfide between lens crystallin and glutathione has been observed in human cataracts and could be formed in vitro by thiol-disulfide exchange reaction. The glutathionyl crystallins have been reported to become partially unfolded. The present paper reports the conformational destabilization by the mixed disulfide formation in calf alpha- and gamma-II crystallin. The conformational stability was studied by the denaturants urea and guanidine hydrochloride (Gdn-HCl), and by proteolytic degradation. The denaturation curves of both urea and Gdn-HCl shift to lower denaturant concentration for crystallins of glutathione mixed disulfide. The decrease in conformational stability is estimated to be 0.22- and 0.92 kcal mol-1 for modified alpha- and gamma-II crystallin, respectively. Proteolytic digestion also shows a faster rate of degradation for the modified crystallins. These results indicate that mixed disulfide destabilizes the crystallin conformation. The destabilization may make crystallins more susceptible to changes as observed in aging lenses.     

Purification and chemical modification of porcine bone morphogenetic protein

Implantation of porcine bone morphogenetic protein (pBMP) in the muscle induces differentiation of mesenchymal-type cells and results in endochondral bone formation. pBMP was isolated from porcine demineralized bone matrix and purified by hydroxyapatite chromatography, Sephadex G75 gel filtration, preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), preparative isoelectric focusing (IEF), and chromatofocusing fast protein liquid chromatography (FPLC). Porcine BMP has an MW of 26 K and a range of pI from 4.65 to 4.73 determined by SDS-PAGE and IEF, respectively. Reconstitution with the citrate buffer supernatant fraction enables as little as 50 micrograms of the soluble pBMP fractions to induce osteogenesis in an in vivo assay. Chemical modification studies indicate that the osteoinductive potential of the pBMP molecule depends on tyrosine, carboxyl groups, and disulfide bonds and can be increased by modification of sulfhydryl groups. Modification of arginine and tryptophan has no effect on bioactivity. By pepsin-limited proteolysis, fragments of pBMP with an MW of 6-14 K show definite, although reduced, BMP activity.     

Microvascular corrosion casting of normal tissue, transitional mucosa and adenocarcinoma in the human colon.

The microcirculatory architecture of normal tissue, transitional mucosa and adenocarcinoma of the human colon was investigated with microvascular corrosion casting (MVCC) combined with scanning electron microscopy (SEM). The study showed that the capillaries within the normal mucosa were arranged in a regular hexagonal pattern around the mucosal glands and that the microvessels of transitional mucosa mostly had lost the typical hexagonal pattern and become slightly wider in diameter. The microvessels in the tumor periphery were increased in number and disorganized, and presented large variation in morphology with claw-like formations, widened sinuses, diverticula and appendixoid patterns. Microvessels were lacking in the central areas of tumors. These morphological alterations may serve as additional indicators of tumor development.     

Different effects of intracellular and extracellular TEA on voltage-dependent K currents of acutely isolated hippocampal CA1 neurons.

Tetraethylammonium (TEA) effects on K currents were examined on either side of the membrane of hippocampal CA1 neurons by means of whole-cell voltage-clamp recording and intracellular perfusion. Recording media contained ion channel blockers to allow the selective activation of voltage-dependent K currents which consisted of a rapidly decaying component (A-current) and a delayed component. Voltage protocols were applied to separate the A-current from the delayed component. Results show that 10 mM extracellular TEA suppressed 50 +/- 11% (S.D., n = 4) of the delayed current at different levels of depolarization but had little effect on the A-current. In contrast 10 mM TEA applied by intracellular perfusion suppressed the A-current by 42 +/- 10% (S.D., n = 4) in addition to inhibiting the delayed currently 55 +/- 15% (S.D., n = 4). Both the intracellular and extracellular actions of TEA on K currents showed no voltage- nor time-dependency. The results suggest that voltage-dependent transient current (A-current) is mediated through a separate group of ionic channels distinct from those that sustained the delayed current. Furthermore, the asymmetrical effects of intracellular and extracellular TEA on the transient current are similar to those described for the A-current in molluscan neurons. This observation supports the notion that the structure of the ion channel mediating the A-current is closely conserved across different species.     

A comparative study of pepsinogen (PG) and lactic acid dehydrogenase (LDH) activities and isoenzyme patterns in tissues from human primary gastric cancer, gastric cancer xenografts in nude mice and gastric cancer cell lines]

Tissue and cell homogenates were prepared for PG and LDH study from 20 samples of histologically proven gastric cancer (GC), 6 samples of gastric cancer xenografts (THPGC-1) of different passages (GCXG) and cultured cells of 3 different gastric cancer cell lines (GCCL). Normal gastric mucosa (NGM) was also obtained from the resected stomach far distant from the primary tumor and histologically tumor free. The results indicated that the expression of PG isoenzymes was low or absent and the PG activities were significantly decreased in GC, GCXG and GCCL as compared to NGM. The activity of LDH was also significantly increased in GC, GCXG and GCCL. In addition, there was a change in isoenzyme pattern in GC and GCXG in which isoenzyme type M was observed whereas isoenzyme type H was preponderent in NGM. The results show that the human gastric cancer xenograft, THPGC-1, has biological properties very similar to those of the primary tumor suggesting that THPGC-1 is a reliable model for the study of the molecular biology of human gastric cancer.     

Surgical treatment of primary small cell carcinoma of the esophagus. Clinical analysis of 11 cases]

545 cases of esophageal carcinoma were confirmed histologically and by esophagoscopic biopsy between Jan. 1982 and May 1990. Primary small cell carcinoma was identified in 19 cases. Of 11 patients confirmed operatively, 5 had oat cell carcinoma, 4 combined oat cell carcinoma and 2 intermediate cell carcinoma. The mean overall survival period was 13.9 months. The longest survival period was 27 months. The prognosis of primary small cell carcinoma was poorer than that of squamous carcinoma of the esophagus because of its propensity of spread and metastasis. Once the diagnosis of small cell carcinoma was established, surgery was undertaken as early as possible. For patients who had lymph node metastasis at the time of diagnosis, we recommended surgical treatment plus systemic chemotherapy after operation. To increase the resection rate, it is important to do chest CT scan, bronchiofiberscopy and B-type ultrasonography before operation.