A physical map of the human salivary proline-rich protein gene cluster covers over 700 kbp of DNA

By using a linking library, we have experimentally linked, ordered, and spaced four of the six loci that constitute the human salivary proline-rich protein (PRP) multigene family. The methods used for mapping these four PRP genes may be useful in other multigene systems in which no probes unique to each member of genes are available, but in which some enzyme site that occurs only once in each member of the family can be found. The remaining two PRP loci have been provisionally mapped and linked within the gene cluster primarily on the basis of the resulting order giving a simple map. The order of the six loci that most simply accounts for our data is PRB2, PRB1, PRB4, PRH2, PRB3, and PRH1. The PRP gene cluster spans at least 700 kbp on chromosome 12 at p13.2. A scheme for the evolution of the cluster that requires an initial gene duplication followed by three unequal but homologous crossovers is given.     

Studies with insulin and insulin-like growth factor-I show that the increased labeling of phosphatidylinositol-3,4-bisphosphate is not sufficient to elicit the diverse actions of epidermal growth factor on MA-10 Leydig tumor cells

In a recent publication we showed that addition of mouse epidermal growth factor (mEGF) to MA-10 Leydig tumor cells rapidly leads to an increase in the incorporation of [3H]inositol-derived radioactivity into an unusual lipid that was identified as phosphatidylinositol-3,4-bisphosphate (PI-3,4-P2). Other ligands that are known to bind to MA-10 cells, such as hCG and arginine vasopressin, however, did not elicit this effect. Inasmuch as mEGF modulates the differentiated functions of MA-10 cells in a number of ways, our findings raised the possibility that PI-3,4-P2 may be an intracellular mediator of these actions of mEGF. In an attempt to answer this question, we set out to determine if other ligands increase the labeling of PI-3,4-P2 in MA-10 cells prelabeled with [3H]inositol, and if such ligands mimic the diverse biological actions of mEGF on these cells. The experiments presented herein show that insulin, insulin-like growth factor-I, and transforming growth factor-alpha increase the labeling of PI-3,4-P2 in MA-10 cells, but only transforming growth factor-alpha mimics the actions of mEGF on the differentiated functions of MA-10 cells. We conclude that an increase in the labeling of PI-3,4-P2 is not sufficient to elicit these actions of mEGF.     

Paraquat toxicity and pyridine nucleotide coenzyme synthesis: a data correction

The decrease in pyridine nucleotide coenzymes which occurs during poisoning of Escherichia coli by hyperbaric oxygen or paraquat is not due to impairment of nicotinatemononucleotide pyrophosphorylase (carboxylating) [EC 2.4.2.19] as was previously proposed (Brown, O.R. et al. Biochem. Biophys. Res. Commun. 91:982-990; 1979). This was shown directly using extracts of E. coli, prepared after exposure to 1 mM paraquat or 4.2 atmospheres of oxygen. The enzyme also was not impaired in Neurospora crassa by 1 mM paraquat. A naturally-occurring, non-dialyzable inhibitor of the enzyme was found in E. coli extracts. The inhibitor caused the erroneous, low nicotinatemononucleotide pyrophosphorylase (carboxylating) activities previously reported in extracts of E. coli poisoned by paraquat.     

Improved Dot-Blot Hybridization Assay for Large-Scale Detection of Potato Viruses in Crude Potato Tuber Extracts

Abstract A rapid and simple technique has been developed to enhance the sensitivity of virus detection in dot-blot hybridization assay by up to 1000 fold. The procedure generally follows that of B aulcombe et al. (1984) but includes moderate heating of the nitrocellulose filter in 10XSSC, 0.5% SDS solution at 55°C after sample application. Using this procedure, four potato viruses (PVX, PVS, PVM and PVY) were detected with cloned virus-specific ³²P-cDNA probes in 2 μl spots containing 0.2–2 pg of purufied virus (0.1–1 ng/ml). The procedure was also successfully applied for the detection of PVX, PVS, PVM and PVY in crude potato tuber extracts.     

The influence of cell interactions and tissue mass on differentiation of sea urchin mesomeres

The developmental potential of different blastomeres of the sea urchin embryo was re-examined. We have employed a new method to isolate substantial numbers of different kinds of blastomeres from 16-cell-stage embryos, and we have used newly available molecular markers to analyze possible vegetal differentiation. We have found that, while isolated mesomere pairs behave according to the classical expectations and develop into ectodermal vesicles, there is a clear effect of reaggregating two or more mesomere pairs. They survive better in long-term culture and, after prolonged periods, they display an astonishing ability to express vegetal differentiation. We also combined mesomeres with stained micromeres or macromeres from the vegetal hemisphere. Although induction of guts and spicules was observed, there was little if any effect of varying the ratio of different blastomeres on the kinds of differentiation obtained.     

Effects of hydrocortisone and aminophylline on plasma leukotriene C4 levels in patients during an asthmatic attack

To study the role of leukotriene C4(LTC4) and the effect of hydrocortisone and aminophylline on plasma LTC4 levels in patients with asthmatic attacks, we measured LTC4 in plasma of 18 asthmatics during a wheezing attack and of 7 normal subjects. Blood samples were obtained before and after treatment with aminophylline and/or hydrocortisone injections. We extracted LTC4 using a Sep-Pak C18 cartridge for the measurement of LTC4 by radioimmunoassay. The plasma levels of immunoreactive LTC4 (i-LTC4) of the normal subjects were 142 +/- 25 pg/ml (n = 7), while those of nonatopic type asthmatic patients with wheezing attacks were 208 +/- 68 pg/ml (n = 15) (p less than 0.01). Before and after treatment with both hydrocortisone succinate (100 mg) and aminophylline (250 mg), 6 asthmatic patients with wheezing attacks had a mean plasma level of i-LTC4 181 +/- 24 and 132 +/- 18 pg/ml (p less than 0.01), respectively. On the other hand, the treatment with aminophylline 250 mg alone increased the i-LTC4 levels from 178 +/- 19 pg/mg to 213 +/- 16 pg/mg (n = 6)(p less than 0.05), while treatment with hydrocortisone succinate 100 mg decreased the i-LTC4 level 0.05 from 284 +/- 99 pg/ml to 249 +/- 85 pg/ml (n = 4)(p less than 0.05). In conclusion, the present study shows that the i-LTC4 level in venous blood of patients with asthmatic attacks is decreased significantly by treatment with hydrocortisone succinate.     

Low dose oral alpha-interferon therapy for patients seropositive for human immunodeficiency virus type-1 (HIV-1)

Thirty eight symptomatic and two asymptomatic patients seropositive for human immunodeficiency virus type-1 (HIV-1) were treated with a natural human interferon alpha (HuIFN alpha). Patients were given 2 IU/kg HuIFN alpha orally once daily in powdered maltose held in the mouth to promote mucosal absorption. This oral immunomodulating HuIFN alpha therapy resulted in an increase in CD4+ lymphocytes, an increase in weight, and a dramatic alleviation of clinical symptoms related to HIV-1 infection.     

Microenvironment created by stromal cells is essential for a rapid expansion of erythroid cells in mouse fetal liver

Mouse stromal cell lines (FLS lines), established from the livers of 13-day gestation mouse fetus, supported the proliferation and differentiation of the erythroid progenitor cells from mouse fetal livers and bone marrow in a semisolid medium in the presence of erythropoietin. A large erythroid colony of over 1000 benzidine-positive erythroid cells was developed from a single erythroid progenitor cell on the FLS cell layer after 4 days of culture. When in close contact with the layer, the erythroid progenitor cells divided rapidly with an average generation time of 9.6 h and mature erythroid cells, including enucleated erythrocytes, were produced. The present studies demonstrate that the microenvironment created by the stromal cells can support the rapid expansion of erythropoietic cell population in the fetal liver of mice.     

Extraction and quantitation of astaxanthin from Phaffia rhodozyma

Summary The rapid, quantitative release of astaxanthin and other carotenoids from the yeast Phaffia rhodozyma is described. Hashed cells are ruptured with dimethylsulfoxide (DMSO) and carotenoids extracted into an organic solvent. Extraction and spectrophotometric quantitation of total carotenoids is rapid, reproducible and only small volumes (0.1–2 ml) of culture are required. HPLC analysis in normal phase silica gel column indicates that astaxanthin comprises 65–95% of the total pigmented carotenoids of P. rhodozyma.