Movements and home range of a common species of tree–shrew,Tupaia glis,surrounding houses of otoacariasis cases in Kuantan,Pahang, Malaysia

ObjectiveTo document movement patterns, home range, nesting behaviour and social organization of 5 individuals (3 males and 2 females) of a common species of tree-shrew, Tupaia glis (T. glis) surrounding houses of otoacariasis cases.MethodsEach shrew was fitted with a transmitter chip radio-collar which operates between the frequencies of 154.13 MHz to 154.21 MHz. Each transmitter was then tracked with a Portable Telemetry Receiver (Sirtrack, New Zealand) fitted with a 3-element Yagi antenna. Collared shrews were located using standard methods of ground-based triangulation. Each location was taken from at least 2 directional fixes and a minimum of 3 compass bearings. Fixes were taken hourly for each collared individual from the time of emergence from nest (beginning of activity) till time of entry into the nest (end of activity) every day for 5 to 7 continuous days. Three series of radio telemetry observations were carried out. The bearings, time and positions of an observer were recorded and later plotted on a graph paper in order to derive coordinates of the collared animal. [These coordinates then analyzed using Ecological Software Solutions (Biotas Version 1.03)].ResultsNests were found in a jack fruit tree, long bushes, and 2 houses. Daily telemetry detections demonstrated 2 individuals of different sex having nests (or a nest) in the same house. All shrews emerged from and returned to their nests between 0601 to 0659 hours and 1901 to 1959 hours, respectively. Both the time of exit from and entry into nest were the same between sexes (P>0.05). Their average total active period was 4.90 to 7.00 hours with a total daily travel distant of 270 m to 382 m. A male and a female shrew can move as far as 3 285 m and 4 591 m, respectively. Active movements of T. glis were during daytime. They regularly entered some houses in the area during day and night except for one individual which visited during daytime only. The sizes of home range and core area for the shrews were 2.00–3.40 ha and 0.05–0.42 ha, respectively. Generally, the mean home range size of females was 20.8% larger than that of males. Females covered a 15.4% slightly higher daily movement range compared to males.ConclusionsThis is the first radio telemetry study in Malaysia to monitor movements and home range of shrews carrying ticks on their body. It demonstrates that shrews are potential carriers of ticks from wild into the houses and their compounds based on their total active periods spent moving around from fruit orchards, secondary forest, plantations and other vegetations to trees in compound of 4 to 7 houses and vice versa. There are also evidences showing shrews have close contact with humans.     

大鼠海马神经细胞体外缺糖缺氧模型制备方法的改进

目的:改进培养大鼠海马神经细胞体外缺糖缺氧模型制备方法,并通过兴奋性氨基酸N-甲基-D-天冬氨酸受体拮抗剂进行验证。方法:实验于2004-09/2005-06在南方医科大学基础医学院神经生物学教研室进行。实验材料:出生1d内清洁级SD大鼠,由南方医科大学实验动物中心提供(合格证号为粤证监字2004B023号)。N-甲基-D-天冬氨酸受体拮抗剂5-甲基二氢丙环庚烯亚胺马来酸(MK-801)和D-2-氨基-5-磷酰基戊酸(d-APV)购自Sigma公司。实验方法:取新生1dSD大鼠海马组织作神经细胞分散细胞原代培养,培养到13d时进行氧/葡萄糖剥夺模型的制备:将neurobasal培养基更换为不含葡萄糖的BSSo培养基,连续充以50mL/LCO2 950mL/LN2(体积比)混合气体。缺氧30,45,60,90min后取出细胞,更换为正常neurobasal培养基,恢复正常条件继续培养。将神经细胞随机分为正常对照组、单纯缺氧组、无糖缺氧组、MK-801组和d-APV组。将10μmol/LMK-801和500μmol/Ld-APV在通50mL/LCO2 950mL/LN2混合气前加入到BSSo培养基,缺氧结束后随BSSo培养基一起去掉。复氧24h后采用MTT比色法测神经细胞成活率及Hoechst荧光染料法测神经细胞凋亡率。结果:①随着氧/葡萄糖剥夺时间延长,神经细胞存活率下降。氧/葡萄糖剥夺30,45,60,90min再复氧24h,神经细胞存活率分别为(81.48±3.84)%、(63.14±3.14)%、(41.73±2.97)%和(16.78±2.12)%。②N-甲基-D-天冬氨酸受体拮抗剂MK-801(10μmol/L)和d-APV(500μmol/L)均能明显增加神经细胞存活率(P<0.05),并且两组细胞存活率与正常对照组比较,差异无统计学意义(P>0.05)。结论:实验制备的海马神经细胞短时间氧/葡萄糖剥夺模型可对神经细胞造成迟发性死亡,兴奋性氨基酸N-甲基-D-天冬氨酸受体拮抗剂可保护氧/葡萄糖剥夺诱导的神经细胞损伤,说明本实验建立的缺氧模型有效并简便可靠,并且缩短了缺氧时间。     

Prostanoids, ornithine decarboxylase, and polyamines in primary chemoprevention of familial adenomatous polyposis

BACKGROUND & AIMS: Familial adenomatous polyposis because of germline mutation of the adenomatous polyposis coli gene is characterized by development of colorectal adenomas and, ultimately, colorectal cancer. The usefulness of colorectal mucosal compounds to predict the effect on adenoma development of primary chemoprevention with the nonsteroidal anti-inflammatory drug sulindac was evaluated. METHODS: A randomized, double-blind, placebo-controlled study of 41 subjects genotypically affected with familial adenomatous polyposis but phenotypically unaffected was conducted. Patients received either sulindac or placebo for 48 months, and development of new adenomas was evaluated. The levels of 5 prostanoids, ornithine decarboxylase, and polyamines were measured serially in normal-appearing rectal mucosa. RESULTS: There were no statistically significant differences between treatment groups in baseline levels of prostanoids, ornithine decarboxylase, or polyamines. At conclusion of the study, 4 of 5 prostaglandin levels were statistically significantly lower in the sulindac group than in the placebo group. Among the subset of patients taking sulindac, 3 of 5 prostaglandin levels were statistically significantly lower in patients who were polyp free than in those who developed polyps. By contrast, there were no statistically significant differences in ornithine decarboxylase or polyamines between treatment groups or in those on sulindac who were polyp free compared with those who developed polyps. CONCLUSIONS: Colorectal mucosal prostaglandin levels, but not ornithine decarboxylase or polyamines, may be valuable biomarkers to assess appropriate drug dosage and medication compliance in patients undergoing primary chemoprevention therapy with sulindac. Reduction of mucosal prostaglandin levels may be necessary to achieve chemopreventive benefit from this agent.     

Establishment and characterization of a human myeloid cell line from Philadelphia chromosome-negative myeloblastic leukemia arising in a patient with myelodysplastic syndrome

A new hematopoietic cell line derived from a patient with Philadelphia chromosome (Ph1)-negative myeloblastic leukemia arising from a form of myelodysplastic syndrome (MDS) is described. This cell line, designated TMM, consists of immature cells with the morphological characteristics of young myeloblasts and grows in suspension culture with a doubling time of about 30 hours. By cytochemical analysis the cultured cells were positive for acid phosphatase. They were free of the Epstein-Barr virus-associated nuclear antigen as well as terminal deoxynucleotidyl transferase. Further phenotypic analysis revealed the expression of the myelomonocytic-specific antigen Leu-M1 and receptors for the Fc portion of IgG. Partial differentiation of these cells could be induced by dimethyl sulfoxide, tetradecanoyl phorbol acetate, or hypoxanthine and resulted in cells of the myeloid series expressing lysozyme and receptors for the C3b complement protein. The karyotype was 46,XY, lacked the Ph1 chromosome, and displayed no abnormalities at the light microscopic level. No rearrangement of the bcr-c-abl gene complex was found. This cell line should be useful for studying an important type of the heterogeneous population constituting Ph1-negative myeloblastic leukemia, arising in this instance from MDS, as well as for studying differentiation and proliferation of human pluripotent stem cells.     

A K-ras oncogene increases resistance to sulindac-induced apoptosis in rat enterocytes

BACKGROUND & AIMS: Mutations of c-K-ras occur commonly in colonic neoplasms. The aim of this study was to determine how c-K-ras mutations alter the responses to the chemopreventive agent sulindac. METHODS: The parental rat intestinal cell line IEC-18 and c-K-ras-transformed derivatives were treated with sulindac sulfide. Cell cycle distribution was determined by flow-cytometric analysis (fluorescence-activated cell sorter), apoptosis by DNA fragmentation (laddering), flow cytometry, and microscopy, and changes in gene expression by immunoblotting. RESULTS: Sulindac sulfide inhibited cell growth and induced apoptosis in a time- and dose-dependent manner more rapidly in and at lower concentrations in parental cells than ras-transformed cells. Expression of the sulindac sulfide arrested cells in G0/G1, but cells entered apoptosis throughout the cell cycle. Proapoptotic protein Bak was relatively high in untreated parental cells and increased markedly after sulindac sulfide but was low in untreated ras-transformed cells and did not increase after sulindac sulfide. Expression of other Bcl-2 family members was unchanged after sulindac sulfide. However, sulindac sulfide reduced levels of cyclin D1 protein and cyclin E- and cyclin D1- associated kinase activity. CONCLUSIONS: c-K-ras-transformed enterocytes are relatively resistant to sulindac sulfide-induced growth inhibition and apoptosis, which may result from specific reduction of bak expression. (Gastroenterology 1997 Dec;113(6):1892-900)     

Southern blot patterns, frequencies, and junctional diversity of T-cell receptor-delta gene rearrangements in acute lymphoblastic leukemia

Southern blot analysis of T-cell receptor (TCR)-delta gene rearrangements is useful for diagnostic studies on the clonality of lymphoproliferative diseases. We have developed 18 new TCR-delta gene probes by use of the polymerase chain reaction (PCR) techniques. Application of these probes for detailed analysis of the TCR-delta genes in normal control samples, 138 T-cell acute lymphoblastic leukemias (T-ALL), and 91 precursor B-ALL allowed us to determine the TCR-delta gene restriction map for five restriction enzymes, as well as the Southern blot restriction enzyme patterns of all theoretically possible TCR-delta gene rearrangements. Based on this information, it appeared that 97% of all 213 detected TCR-delta gene rearrangements in our series of ALL could be detected by use of the TCRDJ1 probe and that the majority (76%) of the 213 rearrangements could be identified precisely. In T-ALL, we found a strong preference for the complete rearrangements V delta 1-J delta 1 (33%), V delta 2-J delta 1 (10%), and V delta 3-J delta 1 (7%) and the incomplete rearrangement D delta 2- J delta 1 (11%). In precursor B-ALL, the majority of rearrangements consisted of V delta 2-D delta 3 (72%) and D delta 2-D delta 3 (10%). The junctional diversity of these 6 preferential TCR-delta rearrangements was analyzed and showed an extensive junctional insertion (approximately 30 nucleotides) for complete V delta-J delta rearrangements, whereas incomplete rearrangements had correspondingly smaller junctional regions. The detailed TCR-delta gene restriction map and probes presented here, in combination with the Southern blot patterns of TCR-delta gene rearrangements, are important for TCR-delta gene studies in ALL; all TCR-delta gene rearrangements can be detected and the majority can be identified precisely. Identification of rearrangements is a prerequisite for subsequent PCR analysis of TCR- delta gene junctional regions, eg, for detection of minimal residual disease during follow-up of ALL patients.     

Overexpression of the major vault transporter protein lung-resistance protein predicts treatment outcome in acute myeloid leukemia

The monoclonal antibody LRP56 recognizes a 110-kD major vault protein (lung-resistance protein [LRP]) overexpressed in several P-glycoprotein- negative (Pgp-), multidrug resistant tumor cell lines. To determine the frequency of LRP overexpression, its prognostic significance, and its relation to Pgp, we analyzed bone marrow specimens from 87 consecutive patients with acute leukemia. Diagnoses included de novo acute myeloid leukemia (AML; 21 patients), leukemia arising from an antecedent hematologic disorder or prior cytotoxic therapy (secondary AML; 27 patients), AML in relapse (29 patients), and blast phase of chronic myeloid leukemia (CML-BP; 10 patients). A granular cytoplasmic staining pattern was detected by immunocytochemistry in 32 (37%) cases, including 7 (33%) de novo AML, 13 (48%) secondary AML, 11 (38%) relapsed AML, and 1 of 10 CML-BP. Among 66 evaluable patients with AML, LRP overexpression was associated with an inferior response to induction chemotherapy (P = .0017). Remissions were achieved in 35% of LRP+ patients as compared with 68% of LRP- patients. Although Pgp adversely affected response in univariate analysis (P = .0414), only LRP had independent prognostic significance when compared in a logistic regression model (P = .0046). Differences in remission duration (P = .075) and overall survival (P = .058) approached significance only for LRP. Sequential specimens from remitting patients receiving treatment with the Pgp modulator cyclosporin-A showed emergence of the LRP phenotype despite a decrease or loss of Pgp at the time of treatment failure (P =.0304). Significant associations were observed between LRP and age greater than 55 years (P = .017), Pgp (P = .040), and prior treatment with mitoxantrone (P = .020) but not with CD34. These findings indicate that overexpression of the novel transporter protein LRP is an important predictor of treatment outcome in AML.     

Clonal circulating cells are common in plasma cell proliferative disorders: a comparison of monoclonal gammopathy of undetermined significance, smoldering multiple myeloma, and active myeloma

The blood of most patients with active multiple myeloma (MM) contains cells related to the bone marrow tumor. However, identifying clonal cells in the blood of patients with monoclonal gammopathy of undetermined significance (MGUS) has been difficult. In this study, we analyzed blood mononuclear cells (BMNCs) from 16 patients with MGUS, 2 with amyloidosis, 8 with smoldering MM (SMM), 2 with indolent MM (IMM), and 15 with active MM using three different methods to detect and quantitate clonal cells, ie, immunofluorescence microscopy (IM) for monoclonal plasma cells, three-color flow cytometry (FC) for CD38(+)CD45- CD45(dim) cells, and the allele-specific oligonucleotide polymerase chain reaction (ASO-PCR). Using ASO-PCR, we were able to detect clonal cells in the blood in 13 of 16 patients with MGUS, 2 of 2 with amyloid, 6 of 8 with SMM, 2 of 2 with IMM, and 13 of 15 with MM. In 9 of the 13 patients with MGUS with blood involvement, the number of clonal cells was very small ( < 0.04% of the BMNCs). The median percentage of clonal cells as determined by ASO-PCR was 0.02 for MGUS, 0.02 for SMM, and 0.24 for MM. Clonal plasma cells or CD38+CD45- CD45(dim) cells were identified by IM or FC in 6 of 16 MGUS patients, 4 of 8 with SMM, and 11 of 15 with MM. In all cases in which IM or FC detected clonal cells, the ASO-PCR was positive. This study shows that, by using ASO-PCR, clonal cells can be found at very low levels in the blood in most patients with MGUS. However, the number of clonal cells in the blood of MGUS patients is less than those with overt MM (P = .006). In contrast to MGUS, patients with active MM are more likely to have identifiable clonal circulating plasma cells (P = .05).     

Phorbol ester stimulation increases sickle erythrocyte adherence to endothelium: a novel pathway involving alpha 4 beta 1 integrin receptors on sickle reticulocytes and fibronectin

Sickle-cell adherence to endothelium has been hypothesized to initiate or contribute to microvascular occlusion and pain episodes. Adherence involves plasma proteins, endothelial-cell adhesion molecules, and receptors on sickle erythrocytes. It has previously been reported that sickle reticulocytes express the alpha 4 beta 1 integrin receptor and bind to cytokine-activated endothelium via an alpha 4 beta 1/vascular- cell adhesion molecule-1 (VCAM-1) interaction. To elucidate other roles for alpha 4 beta 1 in sickle-cell adherence, the ability of activated alpha 4 beta 1 to promote adhesion to endothelium via a ligand different than VCAM-1 was explored. Adherence assays were performed under dynamic conditions at a shear stress of 1 dyne/cm2. Preincubation of sickle erythrocytes with phorbol 12,13-dibutyrate (PDBu) increased adherence of sickle cells eightfold as compared with untreated sickle cells. Normal erythrocytes, whether treated with PDBu or not, did not adhere to the endothelium. Activating anti-beta 1 antibodies 4B4 and 8A2 also increased the adhesion of sickle, but not normal, red blood cell (RBC) adhesion to endothelium. Anti-alpha 4 antibodies HP1/2 and HP2/1, inhibitory antibody 4B5, or an RGD peptide inhibited sickle-cell adherence induced by PDBu. Additional studies were undertaken to examine if fibronectin, a ligand for activated alpha 4 beta 1, was involved in PDBu-induced sickle erythrocyte adherence. Adherence of PDBu-treated sickle cells was completely inhibited by the CS-1 peptide of fibronectin. Fibronectin was detected on the surface of washed endothelium using an antifibronectin antibody in enzyme-linked immunosorbent assays. Antifibronectin antibody pretreatment of endothelial cells inhibited PDBu-induced adherence by 79% +/- 17%. Incubation of sickle RBCs with exogenous fibronectin after PDBu treatment inhibited adherence 86% +/- 8%. Taken together, these data suggest that endothelial-bound fibronectin mediates adherence of PDBu- treated sickle cells. Interleukin-8 (IL-8), a chemokine released in response to bacterial infection, viral infection, or other injurious agents, and known to activate integrins, also increased adherence of sickle erythrocytes to endothelial cells via fibronectin. This novel adherence pathway involving sickle-cell alpha 4 beta 1 activated by PDBu or IL-8 may therefore be relevant in vivo at vascular sites that produce IL-8 or similar agonists in response to vascular injury or immune activation. These observations describe ways in which inflammation and immune responses cause vasoocclusive complications in sickle-cell disease.     

Artificial surface effect on red blood cells and platelets in laminar shear flow

Red blood cell (RBC) effects on platelet adhesion to a nonbiologic test surface (tetrafluoroethylene propylene copolymer) and platelet aggregation during laminar shear flow for shear rates to 5,680 s-1 (corresponding to shear stress to 200 dyne/cm2) were investigated. Results on hemoglobin (Hb) and adenosine diphosphate (ADP) release from RBCs, percent decrease of single platelets in the bulk, and percent of test surface covered with platelets were obtained in a cone-and-plate (CP) viscometer for samples of whole blood, suspensions of RBC ghosts in platelet-rich plasma (PRP), and suspensions of RBCs in either PRP or platelet-poor plasma. Results obtained over the shear rate range studied for samples of normal hematocrit indicated that low-stress shearing led to ADP and Hb release from intact RBCs; shear-induced release of ADP from RBCs was about twice that of platelets, and of the total ADP released, the ADP released from RBCs contributed about six times that of the platelets to single platelet reduction in the bulk and about twice that of the platelets to platelet adhesion, ie, coverage of the test surface with platelets. Results obtained for various hematocrits showed that above a threshold hematocrit of about 25% to 35% the RBCs (suspended in PRP) had a greater contribution to ADP release, platelet adhesion, and platelet aggregation than the platelets themselves. Single platelet reduction for samples of RBC ghosts suspended in PRP correlated with shear rate level and not with shear stress.