Effect of artificial gastrointestinal fluids on the excystation and metacystic development of Entamoeba invadens

The effect of artificial gastric fluid (AGF), containing 0.5% pepsin and 0.6% hydrochloric acid, pH 1.8, in distilled water, on the excystation and metacystic development of Entamoeba invadens was examined. Excystation, which was assessed by counting the number of metacystic amoebae after inducing excystation, was enhanced by pretreatment of cysts with AGF for 30 to 60 min at 37°C but not 26°C. Longer exposure of cysts to AGF significantly reduced their viability. Significant enhancement of excystation was observed by pretreatment of cysts with distilled water only at 37°C. In addition, 0.6% hydrochloric acid had a comparable enhancing effect on excystation to AGF. Metacystic development, when determined by the number of nuclei in amoeba, was slightly enhanced by pretreatment with AGF. An artificial intestinal fluid (AIF), containing 1% pancreatin, 1% sodium bicarbonate, and 5% ox bile, pH 8.0, in distilled water, had a significant toxic effect on cysts, where 1% pancreatin had neither an enhancing effect on excystation nor a toxic effect on cysts, whereas 5% ox bile had a toxic effect on cysts. Pretreatment of cysts with AGF followed by AIF had a similar toxic effect on cysts to that by AIF only. These results suggest that gastric fluid but not intestinal fluid at 37°C contributes to enhancing excystation for Entamoeba infection.     

Effect of proteasome inhibitors on the growth,encystation, and excystation of <Emphasis Type="Italic">Entamoeba histolytica and Entamoeba invadens</Emphasis>

The effect of three proteasome inhibitors, lactacystin, clasto-lactacystin beta-lactone, and MG-132, on the growth, encystation, and excystation of Entamoeba histolytica and Entamoeba invadens was examined. All of these drugs blocked E. histolytica growth in a concentration-dependent manner; lactacystin was most potent for the inhibition and MG-132 showed the inhibitory effect only at higher concentrations. E. invadens was more resistant to these drugs than E. histolytica. Encystation of E. invadens was also inhibited and was more sensitive to the drugs than was growth. Beta-lactone was the most potent encystation inhibitor. The inhibitory effect of lactacystin and the beta-lactone on encystation was slightly and little abrogated by the removal of the drug, respectively. Multinucleation occurred in E. histolytica trophozoites treated with these drugs, being most marked with lactacystin. In contrast, no multinucleation was observed in E. invadens treated with the drugs. Electron microscopy revealed that the treatment of E. histolytica trophozoites with lactacystin led to an increase in the number of cells with many glycogen granules in the cytoplasm. Lactacystin, beta-lactone and MG-132 had no or little effect on the excystation and metacystic development of E. invadens. These results suggest that proteasome function plays an important role for Entamoeba growth and encystation, but has no obvious effect on excystation or metacystic development.     

Molecular cloning and characterization of the repetitive DNA sequences that comprise the constitutive heterochromatin of the W chromosomes of medaka fishes

Among the medaka fishes of the genus Oryzias, most species have homomorphic sex chromosomes, while some species, such as Oryzias hubbsi and Oryzias javanicus, have heteromorphic ZW sex chromosomes. In this study, a novel family of repetitive sequence was molecularly cloned from O. hubbsi and characterized by chromosome in situ and filter hybridization, respectively. This repetitive element, which we designated as a BstNI family element, localized at heterochromatin regions on the W chromosome, as well as on two pairs of autosomes. Homologous sequences to this element were found only in O. javanicus, which is a sister species of O. hubbsi, suggesting that this repeated element originated in the common ancestor of these two species. However, the intensity of the hybridization signals was lower in O. javanicus than in O. hubbsi, and the chromosomal location of this element in O. javanicus was confined to heterochromatin regions on one pair of autosomes. Thus, we hypothesize that this repetitive element was extensively amplified in the O. hubbsi lineage, especially on its W chromosome, after the separation of the O. javanicus lineage. In addition, we also found the W chromosomal location of the 18S–28S ribosomal RNA genes in both O. hubbsi and O. javanicus. Our previous studies showed no linkage homology of the sex chromosomes in these species, indicating that the RNA genes were shared between W chromosomes of different origins. This situation may be explained by a translocation of the sex-determining region with the ribosomal RNA genes in either species or an independent accumulation of the RNA genes as a convergent process during W chromosome degeneration.     

Interactions between vitreous‐derived cells and vascular endothelial cells in vitreoretinal diseases

Purpose: This study aimed to investigate the roles played by vitreous‐derived cells in the pathogenesis of vitreoretinal vascular diseases. Methods: The vitreous was removed from porcine eyes and small pieces were cultured from which vitreous‐derived cells were isolated. Polymerase chain reaction and ELISA were performed to determine the expression of vascular endothelial growth factor (VEGF) and interleukin 6 (IL‐6) at the mRNA and protein levels, respectively. The viability of human retinal endothelial cells (HRECs) exposed to vitreous‐derived cells was assessed by MTT assay. Results: Expression of the mRNA and protein of VEGF and IL‐6 was increased by exposing the porcine vitreous‐derived cells (PVDCs) to interleukin‐1α (IL‐1α), interleukin‐1β (IL‐1β) and tumour necrosis factor α (TNFα), but not to VEGF or IL‐6. The percentage of living human vascular endothelial cells was increased by including VEGF and IL‐6 in the culture media. The viability of HRECs was affected by co‐culturing them with PVDCs that had been exposed to IL‐1α, IL‐1β, IL‐6, TNFα and VEGF. Conclusions: Porcine vitreous‐derived cells are stimulated by IL‐1α, IL‐1β and TNFα, and produce VEGF and IL‐6, which then enhance the proliferation of vascular endothelial cells. This network, including the cytokines and different types of cells, may contribute to the pathogenesis of proliferative vitreoretinal diseases.     

Feasibility and usefulness of the ‘Distress Screening Program in Ambulatory Care’ in clinical oncology practice

Objective: Although the implementation of routine screening for distress is desirable, doing so is difficult in today's busy clinical oncology practice. We developed the ‘Distress Screening Program in Ambulatory Care’ (DISPAC program) as a practical means of screening for and facilitating the treatment of major depression and adjustment disorders in cancer patients. This study assessed the feasibility and usefulness of the DISPAC program in actual clinical situations. Methods: As part of the DISPAC program, nurses administered a psychological screening measure, the Distress and Impact Thermometer (DIT), to consecutive cancer patients visiting an outpatient clinic in the waiting room. The attending physician then recommended psycho‐oncology service referral to all positively screened patients. We compared the proportion of patients referred to a psycho‐oncology service during the DISPAC period with the usual care period. Results: Of the targeted 491 patients treated during the DISPAC period, 91.9% (451/491) completed the DIT; the results were positive in 37.0% (167/451), recommendations for referrals were given to 93.4% (156/167), and 25.0% (39/156) accepted the referral. Ultimately 5.3% (26/491) of the targeted patients were treated by psycho‐oncology service as having major depression or adjustment disorders, a significantly higher proportion than during the usual care period (0.3%; p<0.001). The nurses required 132±58 s per person to administer the DIT. Conclusions: The DISPAC program is useful for facilitating the care of cancer patients with psychological distress. Nevertheless, the acceptance of referrals by patients and the reduction of the burden placed on nurses are areas requiring improvement. Copyright © 2009 John Wiley & Sons, Ltd.     

The synchronization of chemotherapy to circadian rhythms and irradiation in pre-operative chemoradiation therapy with hyperthermia for local advanced rectal cancer. Abbreviations: 5-FU 5-fluorouracil LV Leucovorin APR abdomino-perineal resection

Purpose: The therapeutic and adverse effects of pre-operative chrono-chemoradiation with local hyperthermia for patients with rectal adenocarcinoma were evaluated.

Materials and methods: Pre-operative radiation therapy of a total dose of 40?Gy (n?=?10) or 50?Gy (n?=?19) on the whole pelvis and hyperthermia once a week during the radiation therapy for 1?h were performed for patients with T2–T4 rectal adenocarcinoma. Chemotherapy consisted of 5-FU (250?mg?m^?2?per?day) and LV (25?mg?m^?2?per?day) administered by continuous infusion in the night for 5 days a week in the second and fourth weeks of radiation.

Results: Grade 3+ toxicities were seen only in two patients (6.9%). A significant down staging was seen in 41.4% of all cases and 52.6% of cases with a radiation dose of 50?Gy. Of the patients who had received surgical resection of a tumour, three (11.1%) had no residue pathologically in the specimen and eight (29.6%) had microscopic lesions.

Conclusions: These results yielded a high response rate with minimal toxicities for advanced low-rectal adenocarcinoma. The administration of 5-FU during the sleeping time before irradiation might have an advantage not only as a chronotherapy but also as a radiation sensitizer.     

Efficacy of transplanting cryo-preserved and encapsulated xenogeneic fetal liver fragment as an auxiliary liver support in 90%-hepatectomized rats

BACKGROUND/AIMS: Xenogeneic-hepatocyte or liver-fragment transplantation could be an attractive clinical option in hepatic surgery for patients with impaired liver function if xenogeneic hepatocytes or liver fragments could be preserved for lengthy periods and if immunoisolation could be more easily achieved. METHODOLOGY: Porcine fetal and adult livers were used as xenogeneic transplants in rats. The grafts were stored frozen for more than one year in liquid nitrogen. After thawing, they were evaluated histologically and for potential function for auxiliary liver support in 90%-hepatectomized rats. The efficacy of microporous polypropylene membrane as a macrocapsule for immunoprotection was also examined. RESULTS: Frozen liver fragments could be preserved in liquid nitrogen for more than one year. Fetal fragments were better able to survive under the given conditions than the adult fragments. Macrocapsules protected the grafts from xenoantibodies. The survival rate of encapsulated fetal liver fragment-transplanted recipients on the seventh day after 90%-hepatectomy was 72%, while transplant recipients of fragments of fetal-liver, adult-liver, and encapsulated adult-liver, were 0, 0, and 0, respectively. CONCLUSIONS: Porcine fetal liver fragments survived longer in liquid nitrogen than did the adult ones. The fragments retained their capacity to provide auxiliary liver support in 90%-hepatectomized rats.