Genetic diversity in the coat protein coding region of eighty-six sugarcane mosaic virus isolates from eight countries,particularly from Cameroon and Congo

Summary.  Fifty-eight sugarcane virus isolates were obtained from leaves showing mosaic symptoms, and collected in Cameroon (26 isolates), Congo (20 isolates), Egypt (1 isolate), South Africa (3 isolates) and the U.S.A. (8 isolates). All these isolates belonged to Sugarcane mosaic virus (SCMV) based on the amplification product obtained by RT-PCR with SCMV-specific primers. The amplicons (0.9 kb) from the coat protein (CP) coding region were cloned, sequenced and compared to each other as well as to the sequences (GenBank accessions) of 16 SCMV isolates from sugarcane (Australia, South Africa and U.S.A.) and 12 SCMV isolates from maize (Australia, Germany and China). Maximum likelihood and maximum parsimony analyses robustly supported two major monophyletic groups that were correlated with the host of origin: the SCE or sugarcane group that included all isolates from sugarcane and the MZ or maize group that contained all isolates from maize. The 86 virus isolates were distributed in 13 minor phylogenetic groups, four (I–IV) restricted to maize and nine (V–XIII) to sugarcane. A strong correlation was observed between the sugarcane groups and the geographical origin of the SCMV isolates. Each SCMV type strain from sugarcane (A, B, D, E and SC) was distributed in a different phylogenetic group or subgroup. The 26 isolates from Cameroon constituted a relatively homogenous group (group V) whereas the 20 isolates from Congo belonged to two other groups (VI and VII). All the isolates from Cameroon and Congo were different from the SCMV type strains and other strains or isolates studied so far. It appears, therefore, that the population of SCMV from sugarcane in Africa contains virus genotypes that have not yet been described. Received December 10, 2001; accepted July 24, 2002     

Factor VIII inhibitor-bypassing agents act by inducing thrombin generation and can be monitored by a thrombin generation assay

Factor VIII (FVIII)-bypassing agents have complex modes of action but all control bleeding in inhibitor patients by triggering the generation of thrombin. No routine test is available for monitoring this therapy in patients with inhibitors against FVIII. We present an assay that records FEIBA- or FVIIa-mediated changes in thrombin generation (TG) in FVIII inhibitor plasma samples. In plasma samples spiked with FEIBA TG was normalized above 0.4 U/ml, while for recombinant FVIIa (rFVIIa) more than 12.5 microg/ml were required to induce TG in the absence of tissue factor (TF). Addition of TF increased the TG potential of rFVIIa in vitro. This assay seems suitable for monitoring the pharmacokinetics of inhibitor bypassing agents during treatment and possibly for predicting responses to treatment.     

Pneumococcal pneumonia with empyema and hemolytic uremic syndrome in children: report of three cases.

Streptococcus pneumoniae is an uncommon etiological organism in children with hemolytic uremic syndrome (HUS). Patients with S. pneumoniae-associated HUS commonly have a pneumonia or meningitis. Historically, S. pneumoniae-associated HUS usually has a poor clinical outcome. We report 3 pediatric cases of pneumococcal pneumonia-induced HUS. All 3 patients were <2 years old, had an empyema complicating pneumococcal pneumonia, and developed renal failure with oliguria and required peritoneal dialysis for a period of 9 to 26 days. All children received several transfusions of unwashed packed red cells and platelets. All of the patients survived. Of the 3 cases, 2 had a normal renal function at discharge, and 1 had a mild renal impairment at 16-month follow-up. Our report suggests S. pneumoniae-associated HUS remains a rare but severe complication of invasive pneumococcal infection in children. It is important for pediatricians to note that children with pneumococcal pneumonia with severe hematologic and renal dysfunction should be investigated for evidence of S. pneumoniae-associated HUS.     

Process development of an acellular dermal matrix (ADM) for biomedical applications

The object of this study was to compare the extent of decellularization at each critical step of processing porcine skin to produce an acellular dermal matrix (ADM) for biomedical applications. The results demonstrated that the removal of epidermis using treatment with 0.25% trypsin for 18 h and 0.1% sodium dodecyl sulfate (SDS) for 12 h at room temperature was beneficial for the subsequent treatment to remove cells in the dermal structure. Lengthy incubation in 0.25% trypsin (12 h) and then 560 units/l Dispase (12 h) at 25 degrees C of small pieces of porcine skin from which the epidermis had been removed efficiently removed cells and cellular components from the skin. Histological examinations revealed that the epidermis, dermal fibroblasts, and epidermal appendages were completely removed by these treatments, and the basic dermal architecture of collagen bundles was that of a loose meshwork. Examinations by TEM showed that the characteristics of collagen fibers in the ADM were retained after complete removal of cells present under optimal conditions defined in this study. SDS-PAGE and size-exclusion HPLC revealed that collagen fibers in the ADM were mostly type I and showed two typical component peaks identified as oligomers and monomers, respectively.     

Peri- and postnatal developmental toxicity of the fluoroquinolone antibacterial DW-116 in rats.

DW-116 is a fluoroquinolone antibacterial developed by Dong-Wha Pharmaceutical Industry Co. The aim of this study is to determine the potential adverse effects of this chemical on pregnancy, delivery and lactation of dams and on peri- and postnatal development of F1 offspring. The test chemical was orally administered to pregnant rats from day 16 of pregnancy, through parturition and throughout the period of lactation up to weaning (postnatal day 21) at dose levels of 0, 10, 50, or 250 mg/kg/day. The progeny were examined at birth and subsequently to weaning. Mortality, body weight change, physical signs of postnatal development (pinna detachment, incisor eruption, fur development, eye opening, testis descent and vaginal opening) and behavioral function (righting reflex, negative geotaxis, grip-strength, pupillary reflex, acoustic startle response, rotating rod test, open field test and water-filled T-maze test) were evaluated. When the exposed offspring reached maturity (11 weeks old) their reproductive capacity was assessed. Maternal toxicity was observed only in the highest dose group and was limited to decreased food consumption during the late stage of pregnancy. However, this change was not observed during the lactation period. There were no adverse effects on mortality, clinical signs, body weight, necropsy findings, organ weight of dams in any treatment group. No adverse effects on the offspring were seen with the low and middle doses tested, but the highest dose increased postnatal mortality. The number of stillborn was also increased at the highest dose but the difference was not statistically significant. Meanwhile, no treatment-related effects were observed in clinical sign, developmental and behavioral landmarks and necropsy findings at any dose levels tested. There were no treatment-related effects on the mating of the F1 generation and resulting F2 offspring. The results of this study indicate that the peri- and postnatal administration of DW-116 to female rats results in an increase in postnatal mortality at a minimally maternotoxic dose, i.e., 250 mg/kg/day. Under the experimental conditions, the no-observed-adverse-effect level for peri- and postnatal developmental toxicity was considered to be 50 mg/kg/day.     

Ripe fruit of Solanum nigrum L. inhibits cell growth and induces apoptosis in MCF-7 cells.

Solanum nigrum L. (SNL) has been traditionally used as a herbal plant, whose fruit is believed to have anti-tumor properties, although the mechanism for the activity remains to be elucidated. In this study, we prepared an ethanol extract from ripe fruits of SNL and investigated the mechanism involved in its growth-inhibitory effect on MCF-7 human breast cancer cells. Results from proliferation assay using tritium uptake showed that the proliferative capacity of MCF-7 cells was strongly suppressed in the presence of SNL ethanol extract. This was further confirmed through MTT assay and trypan blue exclusion experiments, which showed a very close correlation between the SNL extract concentration and the surviving cell numbers. The SNL extract-mediated suppression of cell growth was verified to be apoptotic, based on the appearance of DNA laddering, increase in DNA fragmentation, and low fluorescence intensity in nuclei after propidium iodide staining of the cells. Furthermore, the SNL extract was revealed to be a potential scavenger of hydroxyl radicals and DPPH radicals rather than superoxide anions. Collectively, our findings suggest that SNL fruit extract could be used as an anti-oxidant and cancer chemo-preventive material.     

Tc99m sulfur colloid scintigraphy for detection of intrathoracic extramedullary hematopoiesis in patient with beta-thalassemia major--a case report

Extramedullary hematopoiesis (EMH) refers to the production of blood cells outside the bone marrow and is a compensatory mechanism for bone marrow dysfunction. A 34 year-old female patient with beta thalassemia major was found to have multiple large, well-circumscribed radiopaque paravertibral mass lesions in chest radiography. Magnetic resonance imaging (MRI) of the thorax disclosed a right upper apical and two lower thoracic paraspinal mass lesions with heterogeneous isointensity on T1-weighted images and hypointensity on T2-weighted images. Because intrathoracic EMH is suspected in our case, which had obvious bone marrow dysfunction, radionuclide bone marrow scintigraphy is helpful in supporting the diagnosis. Tc99m sulfur colloid scintigraphy demonstrated three intense radioactive thoracic paraspinal mass lesions corresponding to the lesions seen on MRI. We believe whole body bone marrow scintigraphy with Tc99m sulfur colloid is the best convenient noninvasive method for supporting the diagnosis of EMH.     

Ocular hypotensive,vasorelaxant and cyclic AMP intermediation activities of clozapine displaying antiglaucoma properties

Clozapine, an atypical antipsychotic agent with multiple receptor antagonist activities, was investigated in vivo and in vitro to discover its effects on intraocular pressure and blood flow, on the contractility of ciliary muscle and nonophthalmologic blood vessels, on calcium concentration in A7r5 smooth muscle cells, and on cyclic AMP intermediation. The adrenergic and muscarinic mechanisms involved in the above effects were also examined. In rabbits, clozapine (0.1, 0.25, and 0.5%) prolonged (IOP) recovery time and inhibited IOP response. Clozapine (0.1 and 0.25%) also produced a significant increase in ocular blood flow in this iris, ciliary, retina, and choroid at 30 to 180 min after drug administration. In isolated guinea pig thoracic aorta, clozapine relaxed phenylephrine (10 μM)- and KCl (75 mM)-induced contractions, the estimated IC₅₀ values being 20.4 ± 3.1 nM and 10.6 ± 1.8 μM, respectively. Clozapine (0.1–100 μM) inhibited phenylephrine (10 μM)-induced influx of Ca²⁺, the estimated IC₅₀ value being 0.4 ± 0.1 nM. In isolated pig eye ciliary muscles, clozapine (1.0–100 μM) inhibited carbachol (10 μM)-induced contractions, the estimated IC₅₀ value being 5.8 ± 1.2 μM. Clozapine (0.1–100 μM) increased cyclic AMP accumulation in pig's ciliary bodies, including ciliary process, ciliary muscle, and trabecular meshwork. Pretreatment with carbachol (100 μM) first decreased, then increased, clozapine-induced cyclic AMP accumulation. Studies of pretreatments with various muscarinic receptor antagonists at 100 μM revealed that pirenzepine significantly enhanced clozapine (100 μM)-induced cyclic AMP accumulations in trabecular meshwork, while 4-DAMP methiodide inhibited it in ciliary bodies, and methoctramine decreased it in ciliary process. The ocular hypotensive effects of clozapine may be mostly related to its muscarinic agonist/antagonist activities and associated cyclic AMP increasing activities, which lead to ciliary muscle relaxation and a possibly associated increase in uveoscleral outflow. Clozapine's ability to increase blood flow and relax vessels may be attributed to its ability to block α-adrenergic and decrease intracellular calcium. IOP is considered the major cause of glaucoma, and compounds which are capable of reducing IOP are considered useful for glaucoma treatment. Based on the results above, clozapine may potentially be important in the development of new antiglaucoma agents. Drug Dev. Res. 44:163–173, 1998. © 1998 Wiley-Liss, Inc.