Apoptosis, necrosis and cell proliferation-inhibition by cyclosporine A in U937 cells (a human monocytic cell line).

The immunosuppressive drug cyclosporine A (CsA) has been used in both organ transplantation and the treatment of autoimmune disorders. However, the drug causes adverse effects in the kidney, liver and nervous system, characterized by cellular loss in the affected area. Apoptosis has been shown to play a role in CsA-induced cytotoxicity. Because permeabilization of the mitochondrial membrane is a common criterion in most apoptotic settings in vertebrate cells, here we evaluated whether CsA causes loss of mitochondrial function in the pathway leading to cellular cytotoxicity. We found that CsA caused a concentration- and time-dependent loss of cell viability in the U937 cell line. Treatment of cells at a dose of 10 microM CsA resulted in G0/G1 arrest with a concurrent decrease in the number of cells in the S and G2/M phases of the cell cycle. In mechanistic studies related to the loss of viability, treating cells with 10 microM CsA for 24 h resulted in both DNA fragmentation and an increase of annexin-V-positive cells. CsA treatment also increased activity of the cysteine protease caspase-3, decreased the mitochondrial membrane potential and induced the release of cytochrome c into the cytosol. Furthermore, CsA treatment increased the number of cells in the sub-G0/G1 peak, indicative of a reduction in DNA, although this increase was not observed when cells were pre-treated with a broad caspase inhibitor. In the study, we also found that a higher dose of CsA induces LDH release when the cells were incubated for a longer period. Taken together, these data suggest that the mode of cell death induced by CsA is dose- and time-dependent. Short-term incubation with lower doses of CsA arrests cell growth; this arrest overlaps with the occurrence of apoptosis and then with necrosis after longer treatment periods with higher doses of CsA.     

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.     

FDG accumulation in aortic walls

A 65-year-old woman with no symptoms underwent whole-body F-18 fluorodeoxyglucose (FDG) positron emission tomography (PET) for cancer screening. FDG accumulation was detected incidentally in her arterial walls, including the aortic wall. She had no history of inflammatory or cardiovascular disease. Although accumulation of FDG is well recognized in the aortic wall when vasculitis is present, this patient showed no symptoms of active vasculitis during the 22-month follow-up period after the PET study. The aortic wall might be a site where FDG accumulates physiologically in elderly persons.     

Comparison of image reconstruction algorithms in myocardial perfusion scintigraphy

The purpose of this study was to compare the clinical utility of two image reconstruction algorithms in myocardial perfusion SPECT (single-photon emission computed tomography): filtered back-projection (FBP) and ordered subset expectation maximization (OSEM). A rest/stress one-day protocol with 99mTc-MIBI or tetrofosmin was performed on 102 consecutive patients who underwent coronary angiography. After SPECT data acquisition, images were reconstructed with FBP and OSEM algorithms. We assessed diagnostic performance (sensitivity, specificity and accuracy) in detecting coronary artery stenosis and evaluated regional tracer uptake with a 4-point scoring system. Although there were no significant differences in diagnostic performance between FBP and OSEM reconstruction, the OSEM method yielded higher uptake in the RCA area than the FBP method by reducing the count-loss artifact due to hepatic uptake of the tracers. In addition, regional uptake in the LCX area was significantly lower in the OSEM image than in the FBP image; this phenomenon was observed mainly in patients with coronary stenosis and/or infarction in the LCX territory. In conclusion, OSEM and FBP offered comparable diagnostic performance in stress myocardial perfusion SPECT. The OSEM method contributed to reduction of the count-loss artifact in inferior and posterior walls and to easy recognition of hypoperfusion in the LCX area.     

Excision of a choledochal cyst and simultaneous hepatic lateral segmentectomy

We treated a 4-year-old girl with a choledochal cyst (CC) with bilateral intrahepatic involvement. A severe stricture between the enormously dilated left intrahepatic bile duct and the dilated common hepatic duct was found; this necessitated prophylactic hepatic lateral segmentectomy together with excision of the CC to avoid possible stone formation in the cystically dilated left intrahepatic duct. The choice of the combined procedures was based upon long-term results of other patients in our experience. This is the first such procedure to be reported.     

Application of positron emission tomography imaging to cancer screening

Whole-body positron emission tomography (PET) with(18)F-fluorodeoxyglucose (FDG) is a diagnostic modality that can noninvasively survey the entire body and sensitively detect various cancers. In this study, we examined the potential application of whole-body PET for cancer screening in asymptomatic individuals. PET was performed in conjunction with conventional examinations including physical examination, laboratory study, ultrasonography and chest computed tomography. Between September 1994 and March 1999, 3165 asymptomatic individuals participated in 5575 screening sessions (2017 men and 1148 women; mean +/- SD age, 52.2+/-10.4 years). Follow-up periods were no less than 10 months. PET results were compared with the screening outcomes. Within 1 year after screening, malignant tumours were discovered in 67 of the 3165 participants (2.1%). PET findings were true-positive in 36 of the 67 cancers (54%). Most of the 36 patients underwent potentially curative surgery; thus a wide variety of cancers were detected by PET at potentially curable stages. However, PET findings were false-negative in 31 of the 67 patients (46%). 14 of these 31 (45%) were of urological origin. FDG PET imaging has the potential to detect a wide variety of cancers at potentially curable stages. However, PET imaging is not suited to screening test of general population because PET examination involves substantial cost.     

Comparison of methotrexate resistance conferred by a mutated dihydrofolate reductase (DHFR) cDNA in two different retroviral vectors

We previously reported the protection of hematopoietic cells from methotrexate (MTX) toxicity using an N2-based double copy vector containing serine 31 (S31)-mutated dihydrofolate reductase (DHFR) (DC/SV6S31). To examine whether the use of SFG-based dicistronic vectors will lead to improvement in gene transfer over the DC/SV6 vector, we compared the protection provided by MTX to NIH3T3 cells and hematopoietic progenitor cells infected with these retroviral constructs containing the S31 variant DHFR cDNA. In NIH3T3 cells, the 50% effective dose values of MTX conferred by the SFG vector were 8-fold higher than those obtained with the DC/SV6 vector. DHFR mRNA levels were 22-fold and 38-fold higher than that seen for the DC/SV6 vector according to Northern blot and real-time polymerase chain reaction analysis, respectively. However, DHFR protein expression and DHFR enzyme activity were only 1.5-fold and 2-fold higher in the SFG vector, respectively, indicating that the mRNA from the SFG vector is translated less efficiently than the mRNA generated from the DC/SV6 vector. Furthermore, the degree of MTX protection conferred by each vector in both mouse and human hematopoietic cells was the same. These results indicate that the in vitro transduction efficiency and transgene expression of human DHFR in hematopoietic progenitor cells is equally conferred by both vectors.     

A case of cardiac sarcoidosis associated with various clinical symptoms and thrombocytopenia]

A 29-year-old man presented with acute onset of high fever, chest pain and dyspnea. Chest X-ray film showed diffuse interstitial shadows, a cavitary lesion in the left upper lung field and cardiomegaly, but no lymphadenopathy. Abdominal CT scan showed hepatosplenomegaly and multiple small low density areas in the liver and spleen. Electrocardiogram demonstrated multifocal ventricular premature beats and ventricular tachycardia. Cardiac catheterization revealed left ventricular aneurysms. Sarcoidosis was confirmed by lung and liver biopsy. Drug therapy of prednisolone and mexiletine resulted in clinical improvement of symptoms, signs and chest X-ray film, but platelet count decreased gradually. It is suggested that an immune mechanism of sarcoidosis may have been the cause of this thrombocytopenia.     

Effect of intracarotid infusion of etoposide with angiotensin II-induced hypertension on the blood-brain barrier and the brain tissue

Summary This study investigated the effects of the intracarotid infusion of etoposide in combination with angiotensin 11 (AT 11)-induced hypertension on the blood-brain barrier (BBB) and brain tissue in rats. Eighty rats were divided into five groups: Group 1, intravenous infusion of AT 11 to increase arterial blood pressure; Group 2, intracarotid infusion of etoposide at 22.5 mg/m² for 10 minutes; Group 3, intracarotid infusion of etoposide at 75.0 mg/m² for 10 minutes; Group 4, intracarotid infusion of etoposide at 75.0 mg/m² for 20 minutes; Group 5, intracarotid infusion of etoposide at 75.0 mg/m² for 10 minutes with AT 11-induced hypertension. Evans blue staining of the brain was used as a monitor of BBB disruption. Mean arterial blood pressure over the experimental period in Group 1 increased from 86.3 ± 1.3 mmHg (mean ± SEM) to 139.0 ± 2.4 mmHg, and Group 5 from 85.9 ± 1.8 mmHg to 137.3 ± 2.4 mmHg. None of the animals in Group 1 and 2 showed any obvious neurological change, while all the animals in Group 3, 4 and 5 exhibited diminished activity as their sole neurological change throughout the course of the experiment. Slight evidence of BBB disruption was seen in only 25% of the animals in Group 1. Significant BBB disruption was found in the animals in Group 2, 3, 4 and 5. No histological change was observed in any animal in Group 1 and 2. Demyelination, cerebral edema, and neuronal change on the infused hemisphere were observed in half of the animals in Group 3, 4 and 5, but no marked difference was observed among the three groups.At II-induced hypertension is unlikely to have an adverse effect by itself on the BBB and brain tissue. However, intracarotid infusion of high-dose etoposide is capable of producing BBB disruption and irreversible neuronal damage by itself irrespective of infusion rate. Modification of BBB and neuropathological change in case of intracarotid infusion of etoposide with AT 11-induced hypertension primarily caused by etoposide itself and not by the AT II-induced hypertension.