Use of 5-fluorouracil to analyze the effect of macrophage inflammatory protein-1 alpha on long-term reconstituting stem cells in vivo

A macrophage-derived inhibitor of early hematopoietic progenitors (colony-forming unit-spleen, CFU-A) called stem cell inhibitor was found to be identical to macrophage inflammatory protein-1 alpha (MIP-1 alpha). We investigated the effect of MIP-1 alpha on the earliest stem cells that sustain long-term hematopoiesis in vivo in a competitive bone marrow repopulation assay. Because long-term reconstituting (LTR) stem cells are normally quiescent, an in vivo model was first developed in which they are triggered to cycle. A first 5-fluorouracil (5-FU) injection was used to eliminate later progenitors, causing the LTR stem cells, which are normally resistant to 5-FU, to enter the cell cycle and become sensitive to a second 5-FU injection administered 5 days later. Human MIP-1 alpha administered from day 0 to 7 was unable to prevent the depletion of the LTR stem cells by the second 5-FU treatment, as observed on day 7 in this model, suggesting that the LTR stem cells were not prevented from being triggered into cycle despite the MIP-1 alpha treatment. However, the MIP-1 alpha protocol used here did substantially decrease the number of more mature hematopoietic progenitors (granulocyte-macrophage colony-forming cells [CFC], burst- forming unit-erythroid, CFCmulti, and preCFCmulti) recovered in the bone marrow shortly after a single 5-FU injection. In vitro, MIP-1 alpha had no inhibitory effect on the ability of these progenitors to form colonies. This study confirms the in vivo inhibitory effect of MIP- 1 alpha on subpopulations of hematopoietic progenitors that are activated in myelodepressed animals. However, MIP-1 alpha had no effect on the long-term reconstituting stem cells in vivo under conditions in which it effectively reduced all later progenitors.     

Protective effect of prophylactic penicillin on splenectomized mice exposed to an aerosolized suspension of type III Streptococcus pneumoniae

Prophylactic penicillin has been recommended for use in asplenic patients and postsplenectomy patients. A laboratory model using aerosolized pneumococci has been devised to test the effectiveness of prophylactic penicillin in a manner analogous to human experience. There is increased mortality, over time, in asplenic mice exposed to aerosolized type III Streptococcus pneumoniae. One hundred twenty-one male Swiss mice (mean weight 26 g) were divided into four groups: splenectomized, sham-operated, splenectomized + penicillin, and sham- operated + penicillin. After 2 wk the four groups were exposed for 30 min to an aerosolized atmosphere of 2.4 x 10(9) colony-forming units of type III S. pneumoniae using a Tri-R model A-42 airborne infection apparatus. Penicillin was given at a daily intramuscular dosage of 40,000 units procaine penicillin G beginning 2 days prior to exposure and continuing through the third day after exposure. The splenectomized and sham-operated mice given penicillin showed significantly lower mortality (p less than 0.001) than mice not given penicillin.     

Infrequent alterations of the RAR alpha gene in acute myelogenous leukemias, retinoic acid-resistant acute promyelocytic leukemias, myelodysplastic syndromes, and cell lines

Retinoids are important regulators of cell growth and differentiation in vitro and in vivo and they exert their biologic activities by binding to nuclear retinoic acid receptors (RARs; alpha, beta, and gamma) and retinoid X receptors (RXRs; alpha, beta, and gamma). All- trans retinoic acid (RA) induces complete remission in patients with acute promyelocytic leukemia (APL) presumably by binding directly to RAR alpha of APL cells. Leukemic blasts from APL patients initially responsive to RA can become resistant to the agent. HL-60 myeloblasts cultured with RA have developed mutations of the ligand-binding region of RAR alpha and have become resistant to RA. Furthermore, insertion of an RAR alpha with an alteration in the ligand-binding region into normal murine bone marrow cells can result in growth factor-dependent immortalization of the early hematopoietic cells. To determine if alterations of the ligand binding domain of RAR alpha might be involved in several malignant hematologic disorders, the mutational status of this region (exons 7, 8, and 9) was examined in 118 samples that included a variety of cell lines and fresh cells from patients with myelodysplastic syndromes (MDS) and acute myeloid leukemias (AML), including 20 APL patients, 5 of whom were resistant to RA and 1 who was refractory to RA at diagnosis, using polymerase chain reaction-single- strand conformational polymorphism (PCR-SSCP) analysis and DNA sequencing. In addition, 7 of the 20 APLs were studied for alterations of the other coding exons of the gene (exons 2 through 6). No mutations of RAR alpha were detected. Although the sensitivity of PCR-SSCP analysis is less than 100%, these findings suggest that alterations of RAR alpha gene are rare and therefore other mechanisms must be involved in the onset of resistance to retinoids and in the lack of differentiation in disorders of the myeloid lineage.     

Effect of low-dose prednisone (with calcium and calcitriol supplementation) on calcium and bone metabolism in healthy volunteers

The administration of moderate to high doses of corticosteroids is associated with bone loss. This probably results from the uncoupling of bone formation (decreased) and bone resorption (unchanged or increased). We examined the effect of low-dose (10 mg/day) prednisone (LDP) and the possible mitigating effects of calcium and 1.25 (OH)2 vitamin D (calcitriol) on calcium and bone metabolism in eight healthy, young male volunteers. The study consisted of four observation periods: in the first period, LDP was prescribed during 1 week; in the second, third and fourth periods, calcium (500 mg/day), calcitriol (0.5 micrograms b.i.d.) and calcium in combination with calcitriol, respectively, were added to LDP. Bone formation was measured by means of serum osteocalcin, carboxy-terminal propeptide of type 1 procollagen (P1CP) and alkaline phosphatase, bone resorption by means of urinary excretion of calcium, hydroxyproline, (free and total) pyridinoline, (free and total) deoxypyridinoline and serum carboxy-terminal cross- linked telopeptide of type 1 collagen (1CTP). Dietary calcium and sodium intake were maintained at a stable level during the entire study period. Treatment with LDP led to a decrease in osteocalcin, P1CP and alkaline phosphatase (all P < 0.01). Urinary excretion of pyridinolines, hydroxyproline and serum 1CTP did not increase, but remained unchanged or slightly reduced (P < 0.05), depending on the time of measurement and the marker of bone resorption. Parathyroid hormone (PTH) (insignificantly) increased during LDP (+19%) and LDP plus calcium (+14%), but decreased during supplementation with calcitriol (-16%) and calcium/calcitriol (-44%; P < 0.01). Urinary excretion of calcium increased during treatment with LDP and calcitriol (P < 0.05) and calcium/calcitriol (P < 0.05). It is concluded that LDP has a negative effect on bone metabolism, since bone formation decreased while bone resorption remained unchanged or decreased slightly. The increase in PTH during LDP could be prevented by calcitriol combined with calcium supplementation.      

PARP inhibition sensitizes childhood high grade glioma, medulloblastoma and ependymoma to radiation

Poly ADP-ribose polymerase (PARP) is a protein involved in single strand break repair. Recently, PARP inhibitors have shown considerable promise in the treatment of several cancers, both in monotherapy and in combination with cytotoxic agents. Synthetic lethal action of PARP inhibitors has been observed in tumors with mutations in double strand break repair pathways. In addition, PARP inhibition potentially enhances sensitivity of tumor cells to DNA damaging agents, including radiotherapy. Aim of this study is to determine the radiosensitizing properties of the PARP inhibitor Olaparib in childhood medulloblastoma, ependymoma and high grade glioma (HGG). Increased PARP1 expression was observed in medulloblastoma, ependymoma and HGG, as compared to non-neoplastic brain tissue. Pediatric high grade glioma, medulloblastoma and ependymoma gene expression profiling revealed that high PARP1 expression is associated with poor prognosis. Cell growth inhibition assays with Olaparib resulted in differential sensitivity, with IC50 values ranging from 1.4 to 8.4 μM, irrespective of tumor type and PARP1 protein expression. Sensitization to radiation was observed in medulloblastoma, ependymoma and HGG cell lines with subcytotoxic concentrations of Olaparib, which coincided with persistence of double strand breaks. Combining PARP inhibitors with radiotherapy in clinical studies in childhood high grade brain tumors may improve therapeutic outcome.     

正常孕妇步行时骨盆与胸廓在水平面上的运动协调模式

背景：妊娠期间,女性发生了一系列的生理的和身体的变化,人们常认为妊娠引起肌肉骨骼系统的一定程度的变化,会影响孕妇的姿势或步态运动。 目的：观测妊娠步行时水平面上骨盆和胸廓的旋转运动,并进行量化研究,比较正常孕妇和正常女性的差异。 方法：选择怀孕20~34周的孕妇志愿者12人,未怀孕妇女13人为对照组。比较孕妇组和对照组的最舒适步行速度、骨盆和胸廓在水平方向上旋转运动的幅度及两者间运动协调性的差异。分别测定孕妇组和对照组的最舒适步行速度和最大的步行速度,分别计算骨盆和胸廓运动的和谐指数,应用相对傅里叶相和偶合力来描述骨盆与胸廓旋转的运动协调特征。 结果与结论：孕妇的最舒适的步行速度呈显著性下降,孕妇组和对照组的Tempa评分、骨盆和胸廓的旋转运动的和谐指数、相对傅里叶相和偶合力差异无显著性意义。实验未发现妊娠对骨盆和胸廓运动的协调有影响。在运动步态方面,孕妇能够适应怀孕引起的变化。减少最舒适步行速度可以使孕妇节约能量消耗。     

Processing of daily and seasonal light information in the mammalian circadian clock

It is necessary for an organism's survival that many physiological functions and behaviours demonstrate daily and seasonal variations. A crucial component for the temporal control in mammals is the circadian clock located in the suprachiasmatic nucleus (SCN) of the hypothalamus. Neurons in the SCN generate a rhythm in electrical activity with a period of about 24h. The SCN receives photic information from photoreceptive ganglion cells in the retina and processes the information, detecting dawn and dusk as well as encoding day-length. Information processing by the SCN is optimized to extract relevant irradiance information and reduce interferences. Neuronal coupling pathways, including GABAergic signalling, are employed to distribute information and synchronize SCN subregions to form a uniform timing signal. Encoding of day-length is manifested in SCN neuronal activity patterns and may be the product of network interactions rather than being based on the single cell.     

Profiling of apoptosis genes allows for clinical stratification of primary nodal diffuse large B-cell lymphomas

Intrinsic resistance of lymphoma cells to apoptosis is a probable mechanism causing chemotherapy resistance and eventual fatal outcome in patients with diffuse large B cell lymphomas (DLBCL). We investigated whether microarray expression profiling of apoptosis related genes predicts clinical outcome in 46 patients with primary nodal DLBCL. Unsupervised cluster analysis using genes involved in apoptosis (n = 246) resulted in three separate DLBCL groups partly overlapping with germinal centre B-lymphocytes versus activated B-cells like phenotype. One group with poor clinical outcome was characterised by high expression levels of pro-and anti-apoptotic genes involved in the intrinsic apoptosis pathway. A second group, also with poor clinical outcome, was characterised by high levels of apoptosis inducing cytotoxic effector genes, possibly reflecting a cellular cytotoxic immune response. The third group showing a favourable outcome was characterised by low expression levels of genes characteristic for both other groups. Our results suggest that chemotherapy refractory DLBCL are characterised either by an intense cellular cytotoxic immune response or by constitutive activation of the intrinsic mediated apoptosis pathway with concomitant downstream inhibition of this apoptosis pathway. Consequently, strategies neutralising the function of apoptosis-inhibiting proteins might be effective as alternative treatment modality in part of chemotherapy refractory DLBCL.     

Expression of cytotoxic proteins by neoplastic T cells in mycosis fungoides increases with progression from plaque stage to tumor stage disease

Granzyme B (GrB) and T-cell-restricted intracellular antigen (TIA-1) are cytotoxic proteins that are specifically expressed by cytotoxic CD4 or CD8 positive T cells and natural killer cells. Recent studies demonstrated frequent expression of GrB and TIA-1 by neoplastic cells in primary cutaneous CD30(+) large T-cell lymphomas and lymphomatoid papulosis but not in CD30(-) large T-cell lymphomas. In the present study, 74 biopsies from 54 patients with mycosis fungoides (MF) were investigated for the expression of GrB and TIA-1 using immunohistochemistry on paraffin sections. Staining of more than 10% of the neoplastic T cells for GrB or TIA-1 was considered positive. All but two follow-up biopsies had been obtained from patients without extracutaneous disease at the time of biopsy. Expression of TIA-1 and GrB was found in 33 (45%) and 14 (19%) of 74 MF biopsies, respectively. Comparison of biopsies from T3NoMo-stage MF (n = 27) and T2NoMo-stage MF (n = 45) showed increased expression of TIA-1 (55 versus 37%) and GrB (33 versus 9%) in T3NoMo-stage MF. Evaluation of multiple sequential biopsies from successive stages of MF also revealed an increase in the GrB/TIA-1 expression with tumor progression in five of eight cases. A clearcut relation between the expression of TIA-1 and/or GrB and the type of skin lesion biopsied was found. Considering all 74 biopsies, expression of TIA-1 and GrB was found in 18 of 50 (35%) and 5 of 50 (10%) patches or plaques, 9 of 16 (55%) and 3 of 16 (20%) tumors without blastic transformation, and 6 of 8 (75%) and 6 of 8 (75%) tumors with blastic transformation (defined as >50% blast cells). Correlation between GrB/TIA-1 expression in first diagnostic biopsies from patches or plaques from 40 patients with T2NoMo-stage MF and clinical follow-up data did not reveal differences in clinical behavior and survival between patients with (n = 14) or without (n = 26) expression of cytotoxic proteins, indicating that MF expressing cytotoxic proteins should not be considered as a separate group.