Yasushi Kasai

Peritoneal washing cytology combined with immunocytochemical staining and detecting mutant K-ras in pancreatic cancer : Comparison of the sensitivity and availability of various methods

Peritoneal metastases are the second most common site of involvement, following the liver, in pancreatic cancer. Thus, we performed peritoneal washing cytology at laparotomy to diagnose accurately the intraperitoneal spread of carcinoma cells to determine the appropriate therapy. Peritoneal washings were collected at laparotomy from 20 Japanese pancreatic carcinoma patients at Nagoya University Hospital between April 1993 and December 1994. From centrifuged deposits, we examined the cytology by three methods as follows. The first method was conventional cytology, including May- Grünwald and Giemsa, Papanicolaou, periodic acid-Schiff, and Alcian blue. The second method was immunocytochemical staining, using antibodies to carbohydrate antigen (CA19–9) and carcinoembryonic antigen. After extracting DNA from the remaining pellet, we studied the last method, detecting K-ras point mutation, by two-step polymerase chain reaction and restriction fragment length polymorphism analysis. In two cases, peritoneal metastases were macroscopically recognized, and the results of all three methods were positive. In the two other cases, where peritoneal dissemination was not macroscopically recognized, the judgments of conventional cytological study and detecting K-ras point mutation were negative. However, a few malignant cells were found by the immunocytochemical staining method. Judging from their clinical course, the positively stained cells were suggestive of malignancy. At present, the immunocytochemical staining method is the most sensitive of these three methods in peritoneal washing cytology. However, preserving DNA is suitable for repeated examination, and a modified method can be applied. If the sensitivity increases, the method of detecting K-ras has the potential to become the standard for peritoneal washing cytology in pancreatic cancer.

Molecular cloning of murine monoclonal anti-idiotypic Fab

Anti-idiotypic antibodies (Ab2) binding to idiotopes on antibodies with various antigen binding specificities (Ab1) are potential regulators of immunity in a variety of diseases, such as autoimmunity, cancer, and viral, bacterial, or parasitic infections. Furthermore, Ab2 are useful probes for the characterization of receptor/ligand interactions. Thus far, Ab2 production has been limited to the isolation of polyclonal Ab2 from immune sera or monoclonal Ab2 from hybridoma supernatants. However, both approaches have produced a limited number of Ab2. As an alternative approach, we demonstrate here the production of Ab2-Fab by using repertoire cloning. Using HIV-1 as a model system, the Ab2-Fab were generated from the spleens of mice immunized with the virus-neutralizing and syncytia-inhibiting anti-HIV-1 monoclonal antibody 0.5 beta. A bacteriophage lambda vector system was used to express a combinatorial library in Escherichia coli. Iodinated 0.5 beta was used to identify 17 Ab2-Fab clones. DNA sequence analysis of five clones revealed three similar kappa and Fd combinations. The Ab2-Fab bound with high affinity (3.5-6.5 x 10(9) liters/mol) specifically to the Ab1 and not to isotype-matched antibodies with unrelated specificities. The three Ab2-Fab probably bind to the same idiotope on the Ab1 as demonstrated in cross-competition binding studies. The Ab2-Fab inhibited binding of the Ab1 to antigen, and therefore, may functionally mimic the epitope defined by the Ab1. Repertoire cloning of Ab2-Fab may facilitate the generation of Ab2 that have potential as modulators of immune responses against various antigens.

Pedicle myocutaneous flaps for reconstruction following total pelvic exenteration of intrapelvic recurrent rectal cancer: report of a case.

Abstract A vast metastatic tumor mass of recurrent rectal carcinoma in the intrapelvic organs is commonly considered unsuitable for total pelvic exenteration (TPE); first, because it is unlikely that it would improve the prognosis and health-related quality of life of the patient, and second, because of the difficulties involved in this surgical technique. However, by using a plastic surgery technique involving reconstruction by filling the pelvic dead space with pedicle myocutaneous flaps (PMF), primary wound closure and extensive resection of the perineum can be achieved, whereby postoperative metastasis may be prevented. We report herein the case of a 71-year-old man found to have local recurrence in the perineum with extensive invasion of the soft tissue as well as adjacent organs, 20 months after abdominal perineal resection for rectal carcinoma. TPE with extensive resection of the perineal soft tissue was performed, followed by perineal reconstruction and packing of the pelvic dead space with PMF, mainly constructed from the gracilis and sartorius muscles of both femurs. His postoperative course was uneventful and he has remained free of local recurrence and symptomatic perineal complaints for 1 year. In this report, we examine the effectiveness of using the gracilis muscle for PMF in intrapelvic and perineal reconstruction after TPE.

Modification of adriamycin pharmacokinetics by direct electric current in rats.

Adriamycin (ADR, doxorubicin), a drug having cardiotoxicity, is electrically charged as a cation in blood. We therefore investigated whether iontophoresis caused by direct electric current (DC; 50 μ A, 90 min) would cause systemic modification of ADR pharmacokinetics. Cathode and anode were placed into a right kidney and muscles of the abdominal wall, respectively, in six Donryu rats. Urinary excretion of ADR, as measured by catheterizing into the right kidney, was significantly higher in the DC group than in the controls (p < 0.05). Both plasmic and renal ADR clearances were significantly higher in the DC group (p <0.005 and p <0.001, respectively). Tissue ADR concentrations were significantly lower in the DC group (heart:p <0.003; liver and lung: both p <0.05). These results suggest that electric therapy might potentially induce modification of ADR pharmacokinetics by iontophoresis, and that the therapy might effectively change ADR concentration both locally and systemically.