Construction of the plasmid PMEX8-HAK1 and random site-directed mutagenesis of human cytosolic adenylate kinase

The pMEX8-hAK1 vector was devised from the pAK plasmid (Kim J. H. et al., 1989, Protein Engineering 5, 379-386), which could directly express human adenylate kinase proteins without recombination and its single strand DNA could be withdrawn with helper phage for random site-directed mutagenesis. The conserved key residues at Lys21, Lys27, and Thr39 were engineered to obtain mutants for kinetic analysis. Three mutants were obtained as K21P, K27R, and T39S, their specific activities were strikingly reduced compared to those of wild type adenylate kinase. This pMEX8-hAK1 will be a powerful tool for site-directed mutagenesis to detect the substrate-enzyme interaction for human adenylate kinase including various other enzymes.     

Physiologic stress responses to laparoscopic cholecystectomy. A comparison of the gasless and pneumoperitoneal procedures

BACKGROUND: Differences in the physiological stress response to pneumoperitoneal (PP) and gasless abdominal wall-lifting (AWL) procedures used for laparoscopic cholecystectomy have not been properly evaluated. METHODS: We compared leukocyte count, interleukin-6 (IL-6) levels, arterial blood gases, creatinine clearance, plasma renin activity, cardiothoracic ratio, and clinical outcome in 27 patients without systemic complications who underwent laparoscopic cholecystectomy, including 11 by AWL and 16 by PP. RESULTS: Transient leukocytosis and high IL-6 levels were observed at POD 1 (postoperative day) in both groups, but both values returned to baseline by POD 2. IL-6 levels correlated significantly with operation time (p < 0.01). Changes in blood gases, creatinine clearance, plasma renin activity, and cardiothoracic ratio were not different for the two groups. The clinical outcome was similar for both groups. CONCLUSIONS: Our results indicate that both PP and AWL are appropriate for patients without serious complications.     

Antioxidative and anti-hydrogen peroxide activities of various herbal teas

Herbal teas, i.e., extracts of herbs, are popular because of their fragrance and antioxidative activity. Since the antioxidative activity comes mainly from polyphenols, total polyphenol concentrations and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activities in herbal teas were measured and compared. Levels of H₂O₂ in the teas were also examined, since the production of H₂O₂ in beverages such as coffee and green tea, has been reported. Only a small amount of H₂O₂ was detected in the herbal teas just after their preparation with hot water. However, H₂O₂ was gradually produced during incubation at 25 °C after extraction with hot water, especially when the teas were incubated in phosphate buffer at pH 7.4. To examine the anti-H₂O₂ activity of herbal teas, various teas were added to a catechin-enriched green tea, which produce much H₂O₂, and they were incubated at 25 °C for one day. Addition of hibiscus and thorn apple tea decreased the production of H₂O₂ in the catechin-enriched green tea, possibly because of a lowering of the pH of the mixture.     

Clinical and pathological significance of numerical aberrations of chromosomes 11 and 17 in colorectal neoplasms

Numerical chromosome aberrations by interphase cytogenetic analysis have been reported in a few samples of colorectal neoplasms. No studies have defined a distinct relationship between these aberrations and clinicopathological features. To investigate the chromosome aberrations as a marker of invasiveness or prognosis, we conducted an interphase cytogenetic study using fluorescence in situ hybridization and examined 142 colorectal neoplasms consisting of 15 adenomas and 127 cancers. The target chromosomes were chromosomes 11 and 17. We also evaluated the nuclear DNA content as detected by flow cytometry, analyzed the relationship between the frequency of aneusomy and clinicopathological features, and examined the survival rate in these patients. The loss of chromosome 11 was observed in 31% of adenomas, whereas in cancers DNA aneuploidy was observed in 63% of cases, a gain of chromosome 17 was observed in 63% of cases, and a gain of chromosome 11 was observed in 42% of cases. Numerical chromosome aberrations in diploid DNA were also observed. Increased depth of invasion (>/=T3) and advanced Dukes' stage (>/=B) of malignant tumors were associated with a higher frequency of a gain of chromosome 11 (P < 0.01 and P < 0.05, respectively). Increased depth of invasion (>/=T2) in cancers was associated with a higher frequency of a gain of chromosome 17 (P < 0.05). Multivariate analysis of postoperative survival showed that a loss or gain of chromosome 11 was independently associated with a poor prognosis (P < 0.05). Numerical chromosome aberrations appear prior to the alteration of nuclear DNA content as detected by flow cytometry and influence the progression of colorectal cancers. Aneusomy of chromosome 11 is associated with poor postoperative prognosis of primary colorectal cancers.     

Significance of angiogenic factors in liver metastatic tumors originating from colorectal cancers

We examined the expression of vascular endothelial growth factor (VEGF) and microvessel counts expressed by CD34 staining in 39 patients with primary and 44 patients with metastatic liver tumors of metastatic colorectal carcinoma, and 29 patients with nonmetastatic colorectal carcinoma as control in order to determine their value in the evaluation of prognosis and recurrence after hepatectomy. Microvessel counts in primary colorectal carcinomas of the metastatic group were significantly higher than those in control (P<0.05). Neither factor correlated with any clinicopathological feature of primary or metastatic liver carcinomas. Higher microvessel counts in metastatic liver tumors tended to be associated with a shorter disease-free interval to second recurrence in the remaining liver (P = 0.069) and were significantly associated with poor prognosis after hepatectomy (P<0.05). We conclude that microvessel count is an important marker of liver metastasis and prognosis in patients with colorectal carcinoma treated with hepatectomy.     

Relationship between morphological diversity and AgNORs or cathepsin B expression in colorectal cancers

The biological characteristics associated with the morphological diversity of colorectal cancers were investigated to elucidate the causes of this diversity. We examined the proliferative and infiltrating activity of tumor cells, indicated by the mean number of Ag nucleolar organizer region associated proteins (NORs) per nucleus (MNA) and the immunohistochemical response to cathepsin B(CB), in various morphological types of early and advanced colorectal cancers. We examined 73 colorectal cancers obtained by endoscopic and surgical resection. MNA values for sessile and flat-elevated cancers were greater than the values for pedunculate, subpedunculate, and flat-or-depressed early cancers (sessile, P < 0.05). In advanced cancers invading the muscularis propria, protruding cancers showed significantly higher MNA values than small ulcerative cancers (P < 0.01). CB expression increased significantly with the progression of colorectal cancers (P < 0.01), but was not related to morphological diversity in early and advanced cancers. In both sessile and flat cancers, CB expression was higher in moderately differentiated than in well differentiated adenocarcinomas. These results indicate that, in colorectal cancers, protruding early cancers without stalks and protruding advanced cancers have higher proliferative activity than pedunculate or flat early cancers and small ulcerative advanced cancers, respectively, and that CB expression is not associated with morphological diversity, but with depth of invasion and histological differentiation.     

Numerical aberration of chromosome 17 is correlated with multiple primary cancer in colorectal carcinoma

In order to test the hypothesis that attention deficit hyperactivity disorder (ADHD) is related to deficits in somatosensory processing, 49 ADHD male children and 49 matched controls were tested on a wide range of tactile tasks, and somatosensory evoked potentials (SEP) were also recorded. In addition, parents' and teachers' ratings on the children's typical responses to tactile stimuli were obtained. The results show that the ADHD children were less skilled on suprathreshold, but not on threshold tasks than were the controls. Further, a larger percentage of ADHD children were 'tactile defensive'. Finally, the ADHD children showed larger-than-normal amplitudes of late, but not early components of the SEP. These data suggest that some aspects of somatosensory processing by ADHD children are deficient.     

Acquired amylase production induced by radiotherapy in a myeloma patient

A 55-year-old patient with multiple myeloma (IgG-lambda) diagnosed in November 1988 was admitted because of bone pain throughout the body. After plasmapheresis and several courses of chemotherapy, a massive tumor of the left thoracic wall involving the rib appeared. Radiotherapy was performed to ameliorate the severe chest pain, after which myelomatous pleural effusion appeared on the left side. The serum, urine and pleural effusion revealed increased activity of amylase of the salivary type. Amylase activity was also detected in the supernatant of myeloma cells cultured from pleural effusion. We reviewed 12 cases of ectopic amylase-producing multiple myeloma. All the cases except one have been reported from Japan, and hyperamylasemia in these cases was detected at diagnosis or during course of the illness. Moreover, cytogenetic analysis of myeloma cells of previous reports revealed structural abnormalities including chromosome 1, near the amylase gene locus. This case also showed t (1; 10) (q 21?; q 26) by examination of 8 metaphase derived from bone marrow. These observations suggested that ectopic amylase production was induced by irradiation to the plasmacytoma of thoracic wall.