Sandy: a new mouse model for platelet storage pool deficiency

Sandy (sdy) is a mouse mutant with diluted pigmentation which recently arose in the DBA/2J strain. Genetic tests indicate it is caused by an autosomal recessive mutation on mouse Chromosome 13 near the cr and Xt genetic loci. This mutation is different genetically and hematologically from previously described mouse pigment mutations with storage pool deficiency (SPD). The sandy mutant has diluted pigmentation in both eyes and fur, is fully viable and has prolonged bleeding times. Platelet serotonin levels are extremely low although ATP dependent acidification activity of platelet organelles appears normal. Also, platelet dense granules are extremely reduced in number when analysed by electron microscopy of unfixed platelets. Platelets have abnormal uptake and flashing of the fluorescent dye mepacrine. Secretion of lysosomal enzymes from kidney and from thrombin-stimulated platelets is depressed 2- and 3-fold, and ceroid pigment is present in kidney. Sandy platelets have a reduced rate of aggregation induced by collagen. The sandy mutant has an unusually severe dense granule defect and thus may be an appropriate model for cases of human Hermansky-Pudlak syndrome with similarly extreme types of SPD. It represents the tenth example of a mouse mutant with simultaneous defects in melanosomes, lysosomes and/or platelet dense granules.     

Molecular markers near two mouse Chromosome 13 genes,muted and pearl,which cause platelet storage pool deficiency (SPD)

The recessive muted (mu) and pearl (pe) mutations on Chromosome (Chr) 13 cause pigment dilution and platelet storage pool deficiency (SPD) in mice. In addition, mu causes inner ear abnormalities and pe has symptoms associated with night blindness. Using an interspecific backcross involving the wild-derived Mus musculus musculus (PWK) stock, we have mapped 33 microsatellite markers and four cDNAs relative to mu, pe, and another recessive mutation, satin (sa). Analyzing a total of 528 backcross offspring, we found tight linkage between the pigment loci and several microsatellite markers (D13Mit87, D13Mit88, D13Mit137 with mu; and D13Mit104, D13Mit160, D13Mit161, and D13Mit169 with pe). These markers should aid the eventual molecular identification of these specific SPD genes.     

Platelet storage pool disease in hybrid rats. F1 fawn-hooded rats derived from crosses with their putative ancestors (Rattus norvegicus)

Fawn-hooded rats have a hemorrhagic disorder known as platelet storage pool deficiency. Parent stocks of Fawn-hooded, Long-Evans, and Wistar (DAB) rats and the progeny of their reciprocal crosses were studied for coat color and percentage 14C-serotonin uptake and secretion by blood platelets. Progeny of Fawn-hooded and Wistar reciprocal crosses were dark- or light-hooded, whereas all reciprocal crosses with the Long-Evans stock were dark-hooded. The light or fawn hood was significantly associated with the reduced serotonin uptake and secretion characteristic of storage pool efficiency.     

A new abnormality of platelet functions. Association of storage pool disease (thrombocytopathia A) with impaired reactivity of platelets to collagen.

Multiple platelet abnormalities were found in a patient with bleeding symptoms. The platelet content of ADP and PF 4 was decreased and the uptake of 14C-serotonin was impaired. The content of acid phosphatase, beta-glucuronidase and beta-N-acetylglucosaminidase was, however, normal and these enzymes were normally released or made available by bovine fibrinogen or ADP. There was no adhesion of platelet to collagen, which also failed to induce reptilase clot retraction, platelet aggregation and release of any of the platelet constituents. The platelets therefore exhibited signs of thrombocytopathy of a combined type with a decreased storage pool as well as a qualitative dysfunction with impaired reactivity to collagen.     

A congenital defect in platelet prostaglandin production associated with impaired hemostasis in storage pool disease

Prostaglandin (PG) production was found to be impaired in platelets from patients with 'storage pool disease', a well characterized hemostatic disorder. This finding adds to the growing body of evidence which implicates prostaglandin synthesis or a similar arachidonate-consuming process in second phase aggregation and the accompanying platelet release reaction. Prostaglandin production is not impaired in platelets from patients with thrombasthenia. It is thus unlikely that aggregation 'per se' is a stimulus for PG production, because thrombasthenic platelets are virtually incapable of aggregation.     

Genetic lipid storage disease with lysosomal acid lipase deficiency in rats

We describe a new animal model of a genetic lipid storage disease analogous to human Wolman's disease. Affected Donryu rats, who inherited the disease in an autosomal recessive mode, manifested marked hepatosplenomegaly, lymph node enlargement, and thickened, dilated intestine. Morphologically, many characteristic foam cells were observed in livers and spleens. No adrenal calcification could be found in affected rats. Biochemical studies on spleen and liver tissues showed massive accumulation of esterified cholesterol and triglycerides, and deficiency of acid lipase for [14C]-cholesteryl oleate. This animal model could contribute greatly to the clarification of the physiological and pathological roles of lysosomal acid lipase in the metabolism of lipoproteins and cholesterol, and of the pathogenesis of atherosclerosis.     

特异性标记EIAV感染性克隆的构建及鉴定

用FLAG^TM或6His对EIAV疫苗株全基因感染性克隆pFD3的S2分子进行分子标记,以建立EIAV疫苗株与野毒株感染鉴别诊断的方法。标记产物pFD3-FLAG和pFD3-HISADD转染驴胎皮细胞(FDD)后收获衍生病毒并用逆转录酶活性检测和PCR方法确定其感染性。在FDD细胞上盲传至第五代后收获细胞培养上清再感染驴单核巨噬细胞(DL),盲传三代后pFD3-FLAGRT酶活性显示弱阳性,未见明显的细胞病变;pFD3-HISADD为强阳性,且细胞病变效应明显,在电镜下可见明显的病毒颗粒。与父本克隆pFD3相比,在细胞水平上二者复制特性有明显的不同。证明在DL细胞上S2基因的完整性是病毒复制很重要的因素。     

Impaired platelet activation in familial high density lipoprotein deficiency (Tangier disease)

ATP binding cassette transporter A1 (ABCA1) is involved in regulation of intracellular lipid trafficking and export of cholesterol from cells to high density lipoproteins. ABCA1 defects cause Tangier disease, a disorder characterized by absence of high density lipoprotein and thrombocytopenia. In the present study we have demonstrated that ABCA1 is expressed in human platelets and that fibrinogen binding and CD62 surface expression in response to collagen and low concentrations of thrombin, but not to ADP, are defective in platelets from Tangier patients and ABCA1-deficient animals. The expression of platelet membrane receptors such as GPVI, alpha2beta1 integrin, and GPIIb/IIIa, the collagen-induced changes in phosphatidylserine and cholesterol distribution, and the collagen-induced signal transduction examined by phosphorylation of LAT and p72syk and by intracellular Ca2+ mobilization were unaltered in Tangier platelets. The electron microscopy of Tangier platelets revealed reduced numbers of dense bodies and the presence of giant granules typically encountered in platelets from Chediak-Higashi syndrome. Further studies demonstrated impaired release of dense body content in platelets from Tangier patients and ABCA1-deficient animals. In addition, Tangier platelets were characterized by defective surface exposure of dense body and lysosomal markers (CD63, LAMP-1, LAMP-2, CD68) during collagen- and thrombin-induced stimulation and by abnormally high lysosomal pH. We conclude that intact ABCA1 function is necessary for proper maturation of dense bodies in platelets. The impaired release of the content of dense bodies may explain the defective activation of Tangier platelets by collagen and low concentrations of thrombin, but not by ADP.     

Association of isolated adrenocorticotropin deficiency with a variety of neuro-somatic abnormalities in congenital facial diplegia (Moebius) syndrome

A male patient with recurring episodes of hypoglycemic attacks was diagnosed as having isolated ACTH deficiency as well as renal glycosuria and ichthyosis vulgaris. In addition, he had facial diplegia and abducens palsy consistent with Moebius syndrome, muscle atrophy with proximal dominancy, high arched palate, hammer toes, and mental retardation. There was electrophysiological evidence of peripheral neuropathy. Muscle biopsy of the deltoid showed mild myofiber atrophy with occasional cylindrical laminated bodies. The association of these disorders has never been reported and it could be coincidental. However, considering the high rate of association of isolated hypogonadotropic hypogonadism and Moebius syndrome with peripheral neuropathy, the present case may indicate a causal relationship between isolated ACTH deficiency and Moebius syndrome, reflecting the disorders in the organ systems derived from a common ectoderm.     

Characterization of a lectin-binding storage protein from pea (Pisum sativum)

From the storage proteins of the pea (Pisum sativum), the fraction which interacts with the pea lectin by the sugar-binding site was studied. By electrophoretical subunit patterns and other criteria, this fraction resembles the group of the 7S storage proteins (vicilins). The fraction was resolved into subunits by micropreparative SDS PAGE. The N-terminal sequences of the individual subunits were determined. Most of these are identical with published vivilin subunit sequences; therefore this lectin-binding fraction belongs to the vicilins. Selected subunits and tryptic fragments were analysed for amino-acid compositions. Though unequivocal assignments to vicilin segments were possible, significant differences could be recognized, in particular in the tryptic fragments.     

Essential fatty acid deficiency and platelet fatty acids of normotensive and genetically hypertensive rats

Termination of pregnancy in missed abortion and intra-uterine fetal death was accomplished using vaginal suppositories of 20 mg PGE₂ in 31 cases and the results were compared with oxytocin induction (with or without estrogen pre-treatment) in 17 cases at the doses routinely used in our hospital. The PG suppositories proved much more superior (96.7%) than oxytocin (47.7%), but induced a higher rate of side effects. The latter were not serious and were generally tolerated by the patients. There was a positive correlation between duration of fetal retention in utero and the induction expulsion time. The over all patient acceptance of the method was quite favourable and the approach appears to be a definite advance towards management of these cases.     

Elastic energy storage in the hopping of kangaroo rats (Dipodomys spectabilis)

Bipedal hopping of kangaroo rats has been studied by making force plate records and X-ray cinematograph films simultaneously. Calculations using the data so obtained, and
anatomical data, show that energy saving by elastic storage is much less important than
in kangaroos.     

Seven day oral supplementation with Cardax TM (disodium disuccinate astaxanthin) provides significant cardioprotection and reduces oxidative stress in rats

In the current study, the improved oral bioavailability of a synthetic astaxanthin derivative (Cardax^TM; disodium disuccinate astaxanthin) was utilized to evaluate its potential effects as a cardioprotective agent after 7-day subchronic oral administration as a feed supplement to Sprague-Dawley rats. Animals received one of two concentrations of Cardax^TM in feed (0.1 and 0.4%; ∼125 and 500 mg/kg/day, respectively) or control feed without drug for 7 days prior to the infarct study carried out on day 8. Thirty minutes of occlusion of the left anterior descending (LAD) coronary artery was followed by 2 h of reperfusion prior to sacrifice, a regimen which resulted in a mean infarct size (IS) as a percentage (%) of the area at risk (AAR; IS/AAR,%) of 61 ± 1.8%. The AAR was quantified by Patent blue dye injection, and IS was determined by triphenyltetrazolium chloride (TTC) staining. Cardax^TM at 0.1 and 0.4% in feed for 7 days resulted in a significant mean reduction in IS/AAR,% to 45 ± 2.0% (26% salvage) and 39 ± 1.5% (36% salvage), respectively. Myocardial levels of free astaxanthin achieved after 7-day supplementation at each of the two concentrations (400 ± 65 nM and 1634 ± 90 nM, respectively) demonstrated excellent solid-tissue target organ loading after oral supplementation. Parallel trends in reduction of plasma levels of multiple lipid peroxidation products with disodium disuccinate astaxanthin supplementation were observed, consistent with the documented in vitro antioxidant mechanism of action. These results extend the potential utility of this compound for cardioprotection to the elective human cardiovascular patient population, for which 7-day oral pre-treatment (as with statins) provides significant reductions in induced periprocedural infarct size.     

Characterization of changes in land cover and carbon storage in Northeastern China: An analysis based on Landsat TM data

We use Landsat TM time series data for the years of 1991/1992, 1995/1996 and 1999/2000 to characterize land-cover change in northeast China. With the information on land-cover change and the density of vegetation and soil carbon, we assess the potential effect of land-cover change on vegetation and soil carbon in this region. Our results show a large decrease of 2.76(104km2 in forest area and a rapid increase of 2.32(104km2 in urban area. Land-cover changes in northeast China have resulted in a potential maximum loss of 273.2 Tg C for the period of 1991-2000, with a net loss of 95.7 Tg C in vegetation and 177.5Tg C in soil. . The conversion of forests into other land-cover types could have potentially resulted in a loss of 254.6 Tg C for the study period, accounting for 68.8% of the total potential carbon loss in the northeast China. To quantify the net effect of land-cover change on carbon storage will require accounting for vegetation regrowth and soil processes. Our results also imply that forest protectionand reforestation are of critical importance to carbon sequestration in China.     

Characterization of changes in land cover and carbon storage in Northeastern China: An analysis based on Landsat TM data

We use Landsat TM time series data for the years of 1991/1992, 1995/1996 and1999/2000 to characterize land-cover change in northeast China. With the information onland-cover change and the density of vegetation and soil carbon, we assess the potential effect of land-cover change on vegetation and soil carbon in this region. Our results show a large decrease of 2.76×10~4km~2 in forest area and a rapid increase of 2.32×10~4km~2 in urban area. Land-cover changes in northeast China have resulted in a potential maximum loss of 273.2 Tg C for the period of 1991-2000, with a net loss of 95.7 Tg C in vegetation and 177.5Tg C in soil. The conversionof forests into other land-cover types could have potentially resulted in a loss of 254.6 Tg C for thestudy period, accounting for 68.8% of the total potential carbon loss in the northeast China. To quantify the net effect of land-cover change on carbon storage will require accounting for vegeta-tion regrowth and soil processes. Our results also imply that forest protection and reforestation are of critical importance to carbon sequestration in China.     

Clinical linoleic acid deficiency in Dahl salt-sensitive (SS/Jr) rats

Male SS/Jr rats were placed on a specially formulated, high-cholesterol, low-sodium diet at 3 weeks of age. Of the 50 animals on the diet, 40 developed skin lesions ranging from focal areas of alopecia to diffuse areas of moist dermatitis on the head, face, ear pinnae, and neck. Similar lesions were noted later in 17 of 36 SS/Jr rats in a second study group. Histopathologic findings from two affected animals revealed diffuse, hyperplastic, ulcerative dermatitis, with bacterial colonies of cocci in superficial crusts, as well as chronic hepatic inflammation with hepatocellular glycogen and sinusoidal macrophage aggregates suggestive of lipidosis. Results of a fatty-acid profile of the affected rats showed serum linoleic acid levels of 931 to 1566 micromol/liter, whereas those for control (SS/Jr) samples ranged from 2711 to 3145 micromol/liter. Dietary analysis of the specially formulated diet showed that it contained only 0.225% linoleic acid, which is below the recommended level of 0.3 to 0.6%. In light of the clinical and dietary findings, a diagnosis of linoleic acid deficiency was made. The food manufacturer revised its dietary formulation to increase the linoleic acid content to 1.05%, and no further cases of dermatitis developed in any subsequent groups of rats maintained under the same study protocol.     

The expression of inducible nitric oxide synthase (iNOS) in the testis and epididymis of rats with a dihydrotestosterone (DHT) deficiency

In our previous studies, we showed that a finasteride-induced DHT deficiency may cause changes in the morphology of the seminiferous epithelium without any morphological alteration of the epididymis. In this study, we demonstrated the constitutive immunoexpression of inducible nitric oxide synthase (iNOS) in the testis and epididymis of Wistar rats treated with finasteride for 28 days (the duration of two cycles of the seminiferous epithelium) and 56 days (the duration of one spermatogenesis). We noted that a 56-day finasteride treatment mainly caused a decrease in the level of circulating DHT, as well as a statistically insignificant decrease in the level of T. The hormone deficiency also led to a change in the iNOS immnoexpression in the testis and epididymis of the finasteride-treated rats. In vitro, DHT did not modify NO production by the epithelial cells of the caput epididymis even when stimulated with LPS and IFNγ, but it did give rise to an increase in NO production by the epithelial cells of the cauda epididymis without the stimulation. DHT did not have a statistically significant influence on estradiol production by cultured, LPS- and IFNγ-stimulated epithelial cells from the caput and cauda epididymis. In conclusion, our data clearly indicates that a finasterideinduced DHT deficiency intensifies the constitutive expression of iNOS in most rat testicular and epididymal cells, so it can be expected that the expression of inducible nitric oxide synthase (iNOS) could be regulated by DHT. On the other hand, the profile of the circulating DHT and T levels strongly suggests that the regulation of constitutive iNOS expression is complex and needs more detailed study.