Cell cycle regulation by the Wee1 Inhibitor PD0166285, Pyrido [2,3-d] pyimidine, in the B16 mouse melanoma cell line

Background  

Wee1 kinase plays a critical role in maintaining G2 arrest through its inhibitory phosphorylation of cdc2. In previous reports, a pyridopyrimidine molecule PD0166285 was identified to inhibit Wee1 activity at nanomolar concentrations. This G2 checkpoint abrogation by PD0166285 was demonstrated to kill cancer cells, there at a toxic highest dose of 0.5 μM in some cell lines for exposure periods of no longer than 6 hours. The deregulated cell cycle progression may have ultimately damaged the cancer cells. We herein report one of the mechanism by which PD0166285 leads to cell death in the B16 mouse melanoma cell line.     

Nonoperative Treatment of Reperforated Duodenal Ulcer: Report of Three Cases

During the period between 1983 and 1998, a total of 58 patients were admitted to the surgical department of Akita Red Cross Hospital with acute duodenal perforation. Of these 58 patients, 16 were treated operatively and 42 were treated nonoperatively. Among the 38 men and 4 women who received nonoperative treatment, 3 developed reperforation. The incidence of reperforation was 7.1% and the mean average interval from the initial treatment until reperforation was 3.5 years. Endoscopic biopsy and/or serum anti-H. pylori IgG measurement revealed Helicobacter pylori infection in all three patients. No serious complications developed during the nonoperative treatment of reperforation in these three patients, and their recovery was uneventful. The hospital stay ranged from 10 to 18 days, with a mean stay of 12 days after the first perforation and from 14 to 18 days, with a mean stay of 15.6 days after the reperforation. Nonoperative treatment proved successful as a life-saving procedure for reperforation of a duodenal ulcer in all three patients. Received: September 7, 1999 / Accepted: May 30, 2000     

An Experimental Study of Atrial Natriuretic Peptide Levels and the Effect of Inhaled Nitric Oxide After Pneumonectomy

(Received for publication on Feb. 24, 1999; accepted on Nov. 11, 1999)     

Rheumatoid arthritis-related antigen 47 kDa (RA-A47) is a product of colligin-2 and acts as a human HSP47

We previously isolated RA-A47, which is recognized as an antigen of rheumatoid arthritis (RA), from a human chondrosarcoma-derived cell line (HCS-2/8). The N-terminal 21-amino-acid sequence of RA-A47 had 81% homology to the deduced amino acid sequence of the human heat-shock protein (HSP) 47 gene, the colligin gene, and 100% homology to that of the colligin-2 gene. Moreover, as is HSP47, RA-A47 was a heat-inducible collagen-binding protein. To further characterize RA-A47, we isolated ra-a47 cDNA from HCS-2/8 cells and human periodontal ligament fibroblast (HPLF) cells. The isolated ra-a47 cDNAs from both cells were almost the same as that of colligin-2. C₅₀₄ and G₅₀₅ in the cDNA sequences of both cells and C₅₉₈ in the cDNA of HCS-2/8 were different from the corresponding bases of colligin-2 cDNA. These differences were also observed in genomic DNA. colligin cDNA was not isolated. To show that the isolated cDNA encodes RA-A47 protein, it was expressed in Cos-7 cells. The produced protein was 47 kDa and was recognized both with RA sera and antirat HSP47 antibody, indicating that it is RA-A47 and has structural similarity to HSP47. These results taken together with our previous findings show that RA-A47 is the putative colligin-2 gene product and behaves as a human HSP47. Although colligin has been considered the human HSP47 gene, failure to detect the colligin gene and its mRNA suggests that colligin does not exist in human cells and that the HSP47 gene is identical with colligin-2, which encodes RA-A47. Received: January 19, 2000 / Accepted: May 1, 2000     

Lymph node metastases identified with mediastinoscopy in a patient with superficial carcinoma of the esophagus

Superficial esophageal cancers limited to the lamina propria are not associated with lymph node metastases. Mediastinoscopic transhiatal esophagectomy was planned in a patient with widespread superficial cancer of the midthoracic esophagus. Sampling of the upper mediastinal lymph nodes revealed metastases. The operation was converted to a transthoracic esophagectomy with radical lymphadenectomy. Histopathologic examination of the resection specimen showed three metastatic lymph nodes, despite local invasion limited to the lamina propria. This is the first report of a patient with superficial esophageal cancer and lymph node metastases.     

Retrodental synovial cyst which disappeared after posterior C1-C2 fusion: A case report

Synovial cysts of the cervical spine are extremely rare. We describe an 8-year-old boy with atlantoaxial subluxation and hypoplasia of the dens. Magnetic resonance imaging showed a round lesion, posterior to the odontoid process. This mass was characterized by a low signal intensity on T1-weighted images, and high signal intensity on T2-weighted images. The retrodental synovial cyst disappeared after posterior atlantoaxial arthrodesis.     

Inducible nitric oxide synthase attenuates endothelium-dependent renal microvascular vasodilation

Previous studies have demonstrated that inducible nitric oxide synthase (iNOS) plays a key pathophysiologic role during sepsis. The present study was designed to delineate the consequences of iNOS activation on renal microvascular function. Male Sprague-Dawley rats were given intraperitoneal injections of lipopolysaccharide (LPS; 4 mg/kg) at 16 h and 4 h before experimentation. Afferent and efferent arteriolar diameters from LPS-treated and control rats were assessed in vitro with the use of the blood perfused juxtamedullary nephron technique. Basal afferent and efferent arteriolar diameters of LPS-treated rats averaged 19.7 +/- 0.9 (n = 7) and 18.3 +/- 1.0 microm (n = 5), respectively, and were similar to those of control rats (20.8 +/- 0.3 [n = 6] and 18.4 +/- 0.6 microm [n = 6], respectively). Superfusion with the selective iNOS inhibitor S,S'-(1,3-phenylenebis[1,2-ethanediyl]) bisisothiourea (PBIT), at the doses of 0.01, 0.1, and 1 microM, significantly decreased afferent and efferent arteriolar diameters in a dose-dependent manner, whereas afferent or efferent arteriolar diameters of control rats were not altered in response to the same doses of PBIT. In the second series of experiments, superfusion with 10 microM acetylcholine (ACh) significantly increased afferent and efferent arteriolar diameters of LPS-treated rats by 14.9 +/- 1.6% (n = 9) and 6.6 +/- 1.1% (n = 6), respectively. The ACh-induced afferent and efferent arteriolar dilator responses were inhibited by superfusion with the nonselective NOS inhibitor N:(omega)-nitro-L-arginine (100 microM). However, afferent and efferent arteriolar dilator responses to ACh were significantly enhanced during selective iNOS inhibition with 1 microM PBIT (40.1 +/- 0.7% and 25.2 +/- 1.3%, respectively). These results suggest that activation of iNOS by LPS increases the influence of nitric oxide on afferent and efferent arteriolar tone and impairs endothelium-dependent nitric oxide effects.     

Multiplexed analysis of post-PCR fluorescence-labeled microsatellite alleles and statistical evaluation of their imbalance in brain tumors.

Detection of the loss of chromosomal regions in cancerous tissues has diagnostic and prognostic relevance, and the development of a reliable and cost-effective technique for this is clinically important. Here we present an efficient technique for quantitative detection of microsatellite alleles, using a post-PCR fluorescence-labeling procedure and multiplexed analysis. We also present a new statistical method for the interpretation of the data that permits reliable and sensitive evaluation of the allelic status of sampled DNA. A high-resolution analysis of allelic imbalance on chromosomes 1p, 10 and 19q in 28 glioma samples of various types using this method revealed that allelic imbalances are more frequent than have been reported, suggesting the diagnostic value of this method in examining the genetic profiles of gliomas.     

The enzymatic basis for the conversion of nonfucosylated to fucosylated alpha-fetoprotein by acyclic retinoid treatment in human hepatoma cells: activation of alpha1-6 fucosyltransferase.

The purpose of the present study was to investigate the mechanism by which nonfucosylated alpha-fetoprotein (AFP) is converted to fucosylated AFP in human hepatoma cell lines exposed to acyclic retinoid (AR), an effective drug for the secondary prevention of hepatocellular carcinoma. AR treatment (100 microM) of HepG2 and Hep3B cells significantly increased the activity and mRNA levels of alpha1-6 fucosyltransferase (alpha1-6 FucT), the enzyme responsible for the fucosylation of AFP, leading to an increase in fucosylated glycoproteins as evidenced by lectin binding measurements. Lectin immunoelectrophoresis of AFP obtained from culture media indicated that the relative percentage of nonfucosylated AFP (L1 fraction) was decreased and alpha1-6 fucosylated AFP (L3 fraction) was increased in these hepatoma cell lines after treatment with AR. The total AFP levels were, however, markedly suppressed by AR treatment, and therefore the absolute L3 fraction on the basis of the total AFP present was extremely low. These results demonstrate that AR enhances the conversion of the L1 to the L3 fraction due to the activation of alpha1-6 FucT in human hepatoma cell lines despite clinical outcome with AR treatment and the L3 fraction of AFP. Even though the dramatic decrease in AFP is the limiting factor in the synthesis of the L3 fraction and, therefore, the absolute value of fucosylated AFP is extremely low, the conversion from L1 to L3 as judged by lectin immunoelectrophoresis represents a good marker for the progress of AR treatment.