Disrupted erythropoietin signalling promotes obesity and alters hypothalamus proopiomelanocortin production.

Although erythropoietin (Epo) is the cytokine known to regulate erythropoiesis, erythropoietin receptor (EpoR) expression and associated activity beyond haematopoietic tissue remain uncertain. Here we show that mice with EpoR expression restricted to haematopoietic tissues (Tg) develop obesity and insulin resistance. Tg-mice exhibit a decrease in energy expenditure and an increase in white fat mass and adipocyte number. Conversely, Epo treatment of wild-type (WT)-mice increases energy expenditure and reduces food intake and fat mass accumulation but shows no effect in body weight of Tg-mice. EpoR is expressed at a high level in white adipose tissue and in the proopiomelanocortin (POMC) neurons of the hypothalamus. Although Epo treatment in WT-mice induces the expression of the polypeptide hormone precursor, POMC, mice lacking EpoR show reduced levels of POMC in the hypothalamus. This study provides the first evidence that mice lacking EpoR in non-haematopoietic tissue become obese and insulin resistant with loss of Epo regulation of energy homeostasis.

Eosinophilic gastrointestinal disorders in infants: a Japanese case series.

BACKGROUND: Eosinophilic gastrointestinal disorders (EGIDs) are disorders characterized by primary eosinophil inflammation in the gastrointestinal tract. There are a small number of reports of eosinophil infiltration in gastrointestinal tracts presenting as EGIDs in infants. In this study, we present Japanese cases of EGIDs in infants. METHODS: Five patients diagnosed with or strongly suspected to have EGIDs in our hospital from 2008 to 2010 were reviewed. Radiographic contrast enema examinations and/or endoscopies were performed in 4 and 3 patients, respectively. RESULTS: There were patients with eosinophilic colitis (1 suspected and 2 biopsy-proven), a patient who was suspected of having allergic eosinophilic enterocolitis, and a patient with eosinophilic gastroenteritis associated with pediatric hypereosinophilic syndrome. CONCLUSIONS: The causes and clinical findings of patients with intestinal eosinophil inflammation vary. Therefore, deliberate examination and observation are important for patients with infantile EGID.

Human pharmacokinetic prediction of UDP-glucuronosyltransferase substrates with an animal scale-up approach.

The aim of the current study was to evaluate the accuracy of allometric scaling methods for drugs metabolized by UDP-glucuronosyltransferases (UGTs), such as ketoprofen, imipramine, lorazepam, levofloxacin, zidovudine, diclofenac, furosemide, raloxifene, gemfibrozil, mycophenolic acid, indomethacin, and telmisartan. Human plasma clearance (CL) predictions were conducted from preclinical in vivo data by using multiple-species allometry with the rule of exponents and single-species allometric scaling (SSS) of mice, rats, monkeys, or dogs. Distribution volume at a steady state (V(ss)) was predicted by multiple-species allometry or SSS of V(ss). Oral plasma clearance (CL(po)) was calculated under the assumption that F(a) × F(g) was equivalent across species. Each of the results was compared with the observed parameter calculated from the clinical data after intravenous or oral administration. Multiple-species allometry and SSS of mice, rats, and dogs resulted in a similar accuracy of CL and CL(po) predictions. Monkeys tended to provide the most accurate predictions of human CL and CL(po). The ability to predict the half-life, which was determined from CL and V(ss) predictions, was more accurate in SSS of rats and monkeys. The in vivo fraction metabolized by glucuronidation (f(m,UGT)) in bile duct-cannulated monkeys was relatively similar to that of humans compared with other animal species, which likely contributed to the highest accuracy of SSS prediction of monkeys. On the basis of the current results, monkeys would be more reliable than other animal species in predicting human pharmacokinetics and f(m,UGT) for drugs metabolized by UGTs.

Cerebral substrates related to impaired performance in the clock-drawing test in dementia with Lewy bodies.

AIMS: To identify the neural correlates of impaired performance in the clock-drawing test (CDT) in patients with dementia with Lewy bodies (DLB). METHODS: Cerebral blood flow was measured by single photon emission computed tomography in patients with clinically diagnosed DLB, and was compared between impaired CDT (n = 30) and normal CDT (n = 30) subgroups. RESULTS: DLB patients with impaired CDT performance showed significantly lower cerebral blood flow in the bilateral frontal eye fields, supplementary eye fields, right posterior putamen and the right ventrolateral part of the thalamus relative to the normal CDT subgroup. Performance in other visuospatial/attentional tasks (trail making test part A, copying a cube, semantic fluency, and block design) was also poorer in the impaired CDT group than the normal CDT group. CONCLUSIONS: This study indicates that impaired performances on the CDT and some visuospatial/attentional tasks by DLB patients are closely related to dysfunctions of the frontal-subcortical network relevant to control of visuospatial attention and arousal, involving the frontal eye fields, supplementary eye fields and the thalamus. Our findings provide evidence that cognitive performance in DLB reflects the pathological involvement of both cortical and subcortical regions, and suggest that the neurophysiological basis underlying impaired CDT in DLB may be different from that in Alzheimer disease.

[Pilot study of primary systemic chemotherapy with docetaxel (DOC), epirubicin (EPI) and capecitabine (Xeloda) in patients with advanced breast cancer].

We evaluated the safety and efficacy of primary systemic chemotherapy (PSC) with docetaxel (DOC), epirubicin (EPI) and capecitabine (Xeloda:XLD) in 10 patients with advanced breast cancer. Their mean age was 54.7 years,and preoperative stages were IIB, seven cases; IIIA, two; and IV, one,respectively. The regimen consisted of XLD (2,400 or 3,000 mg/day) orally for 14 consecutive days, and DOC (60 or 70 mg/m2) and EPI (50 or 60 mg/m2) intravenously on day 8. This was repeated 4 times every 3 weeks. One patient discontinued this regimen after one course at her own request. Although the results revealed leucopenia and neutropenia of more than grade 3 in 8 and 10 patients,they could be treated on an outpatient basis with the use of G-CSF to maintain this regimen. Alopecia of grade 2 was found in all patients,neutropenic fever of more than 38.5 degrees C in 5,and hand-foot syndrome in 3. Downstaging after PSC was demonstrated in 7 cases (Stage IIB to I, three cases; IIB to IIA, three; and III A to I, one), with a response rate of 77.8%. Breast conserving therapy was performed in 8/10 patients. Pathological findings on cytological degeneration showed grade 0, one; grade 1a,seven; grade 2, one; and grade 3, one, respectively. Axillary lymph node metastasis was revealed in 7 cases. This regimen would be an alternative to PSC on an outpatient basis while taking great care of myelosuppression and hand-foot syndrome.

Radiosensitization by hyperthermia in the chicken B-lymphocyte cell line DT40 and its derivatives lacking nonhomologous end joining and/or homologous recombination pathways of DNA double-strand break repair.

Hyperthermia has a radiosensitizing effect, which is one of the most important biological bases for its use in cancer therapy with radiation. Although the mechanism of this effect has not been clarified in molecular terms, possible involvement of either one or both of two major DNA double-strand break (DSB) repair pathways, i.e. nonhomologous end joining (NHEJ) and homologous recombination (HR), has been speculated. To test this possibility, we examined cells of the chicken B-lymphocyte cell line DT40 and its derivatives lacking NHEJ and/or HR: KU70(-/-), DNA-PKcs(-/-/-), RAD54(-/-) and KU70(-/-)/RAD54(-/-). Radiosensitization by hyperthermia could be seen in all of the mutants, including KU70(-/-)/RAD54(-/-), which lacked both NHEJ and HR. Therefore, radiosensitization by hyperthermia cannot be explained simply by its inhibitory effects, if any, on NHEJ and/or HR alone. However, in NHEJ-defective KU70(-/-) and DNA-PKcs(-/-/-), consisting of two subpopulations with distinct radiosensitivity, the radiosensitive subpopulation, which is considered to be cells in G(1) and early S, was not sensitized. Substantial sensitization was seen only in the radioresistant subpopulation, which is considered to be cells in late S and G(2), capable of repairing DSBs through HR. This observation did not exclude possible involvement of NHEJ in G(1) and early S phase and also suggested inhibitory effects of hyperthermia on HR. Thus partial contribution of NHEJ and HR in radiosensitization by hyperthermia, especially that depending on the cell cycle stage, remains to be considered.

Characterization of two types of cytochrome P450 aromatase in the serial-sex changing gobiid fish, Trimma okinawae.

To investigate the role of estrogen in the serial-sex changing fish Trimma okinawae, we isolated complementary DNAs encoding two distinct cytochrome P450 aromatase isoforms from adult ovary and brain (termed P450aromA and P450aromB, respectively). Sequence and phylogenic analyses showed that the goby P450arom forms belong to two separate CYP19 subfamilies. Transient expression of these cDNAs in HEK293 cells caused conversion of exogenous testosterone to estradiol-17beta. RT-PCR showed that P450aromA was expressed in the brain, spleen, testis and ovary. P450aromB was expressed in the brain, liver, testis and ovary. In situ hybridization studies showed that P450aromA mRNA, but not P450aromB mRNA, was present in both ovary and testis. Positive signals were restricted to granulosa cells of vitellogenic follicles and interstitial cells of mature testis. Ovarian expression of both P450arom genes during the spawning cycle was examined by quantitative real-time RT-PCR. P450aromA transcripts increased during vitellogenesis and decreased prior to spawning. In contrast, P450aromB transcripts were barely detectable and did not correlate with ovarian development. These findings suggest that P450aromA, but not P450aromB, is involved in regulating ovarian vitellogenesis in goby.

Encephalopathy in megacystis-microcolon-intestinal hypoperistalsis syndrome patients on long-term total parenteral nutrition possibly due to selenium deficiency.

This report concerns two patients (female, 9 and 6 years) who were diagnosed with megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIHS). Although they exceeded the usual life expectancy of patients diagnosed with MMIHS because of total parenteral nutrition (TPN), they demonstrated progressive neurological deficits and showed histopathological features in the brain. Both patients were diagnosed with MMIHS in the neonatal period and were fed by TPN. The first patient developed visual and gait disturbances at the age of 7 years. Two months later, she developed dysarthria and muscular weakness, and could not maintain her posture. The level of serum selenium was extremely low. The second patient developed flexion and spasticity of the extremities followed by decorticate posture at the age of 3 years. Both patients died of sepsis. The brain weights of the two cases were 880 g and 715 g. In both cases, severe neuronal loss and gliosis were present in the medial convolutions of the occipital lobe, including the visual cortex. The postcentral gyrus and temporal transverse gyrus were also involved. In addition, extensive loss of Purkinje cells and granular neurons, and gliosis were observed in the cerebellum. We measured the selenium content of the brains and livers using the graphite furnace atomic absorption spectrometry method. Selenium was not detected in either brain, although the livers of both cases contained a low level of selenium. On immunohistochemical examination of the anti-oxidative enzymes, histiocyte-macrophage lineage cells in MMIHS cases, including microglia and Kupffer cells, showed only a weak reaction for glutathione peroxidase, of which selenium is an essential component. However, the cells in the control cases were strongly positive. In cases of MMIHS and methylmercury intoxication, the brain features similar lesions, in both their topographical and histopathological aspects. We considered that the brain lesions of the MMIHS patients mainly resulted from oxidative damage of the brain related to the low levels of glutathione peroxidase and other selenoproteins due to selenium deficiency.

SDF-9, a protein tyrosine phosphatase-like molecule, regulates the L3/dauer developmental decision through hormonal signaling in C. elegans.

The dauer larva of the nematode Caenorhabditis elegans is a good model system for investigating the regulation of developmental fates by environmental cues. Here we show that SDF-9, a protein tyrosine phosphatase-like molecule, is involved in the regulation of dauer larva formation. The dauer larva of sdf-9 mutants is different from a normal dauer larva but resembles the dauer-like larva of daf-9 and daf-12 dauer-constitutive mutants. Like these mutants, the dauer-constitutive phenotypes of sdf-9 mutants were greatly enhanced by cholesterol deprivation. Epistasis analyses, together with the relationship between sdf-9 mutations and daf-9 expression, suggested that SDF-9 increases the activity of DAF-9 or helps the execution of the DAF-9 function. SDF-9 was expressed in two head cells in which DAF-9 is expressed. By their position and by genetic mosaic experiments, we identified these cells as XXXL/R cells, which are known as embryonic hypodermal cells and whose function at later stages is unknown. Killing of the sdf-9-expressing cells in the wild-type first-stage larva induced formation of the dauer-like larva. Since this study on SDF-9 and former studies on DAF-9 showed that the functions of these proteins are related to those of steroids, XXXL/R cells seem to play a key role in the metabolism or function of a steroid hormone(s) that acts in dauer regulation.

Quantitative analysis of correlation between number of nuclear plasmids and gene expression activity after transfection with cationic liposomes.

PURPOSE: A quantitative understanding of the intracellular trafficking of plasmids delivered by nonviral vectors is essential for optimizing vector functions to increase their transfection efficiency. In this study, quantitative methods were developed to measure plasmids delivered to the nucleus, and the relationship between transfection activity and the number of plasmids in the nucleus were analyzed. METHODS: AH130 cells were transfected with plasmids in cationic liposomes at various doses. The nuclear fraction was isolated after NP-40 lysis. and the unincorporated plasmids were enzvmatically degraded and washed away. Intranuclear plasmids were amplified by quantitative PCR. and the number of plasmids was determined. Plasmid amounts in the nucleus were also measured by Southern analysis to confirm the quantification. RESULTS: Both methods led to similar results in measuring the nuclear plasmids within the same order of magnitude. A remarkable saturation was found for transfection activity vs. number of plasmids in the nucleus, whereas no saturation was observed in nuclear-delivered plasmids vs. dose. CONCLUSIONS: These results clearly demonstrate the importance of the quantitative measurement of intracellular trafficking of plasmids after transfection. The findings herein described suggest that efficient transgene expression as well as enhanced nuclear delivery is required in order to achieve the maximal transfection activity of nonviral vectors.