Inhibition of tumor metastasis of lewis lung carcinoma in C57BL/6 mice by intrapleural administration of Lactobacillus casei

Summary The antimetastatic effect of Lactobacillus casei YIT9018 (LC 9018) against Lewis lung carcinoma (3LL) in C57BL/6 mice was determined. Intrapleural (i.pl.) administration of LC 9018 was effective in inhibiting pulmonary metastasis after s.c. inoculation of 3LL tumors into C57BL/6 mice. The combination of i.pl. and intralesional or i.v. injections of LC 9018 also markedly inhibited pulmonary metastasis in 3LL-bearing mice. The i.pl. administration of LC 9018 into mice induced an increase in the number of thoracic exudate cells (TEC) and the cell population in the TEC was mainly polymorphonuclear leukocytes in the early stage, while macrophages were dominant in the late stage. In addition, in vitro cytolytic activity against 3LL cells and natural killer cell activity of TEC were augmented by the i.pl. administration of LC 9018. Furthermore, i.pl. administration of LC 9018 into the mice rendered their lung macrophages tumoricidal for 3LL cells in vitro. These results show that TEC induced by i.pl. administration of LC 9018 played a key role in the inhibtion of metastasis in 3LL-bearing mice.     

Hydrogen Peroxide-dependent Synthesis of Flavonols in Mesophyll Cells of Vicia faba L.

Mesophyll cells of Vicia faba contain kaempferol and quercetinglycosides. When isolated mesophyll cells were treated with0.1 mM H₂O₂ for 2 h, the levels of these flavonols increasedby 10–70% of the control values (mean values, 19.6% and34.4% for kaempferol and quercetin glycosides, respectively).Such increases in levels of flavonols were also observed inisolated vacuoles of mesophyll cells. However, when mesophyllcells and vacuoles were treated with 10 mM H₂O₂₎degradationof flavonols was observed. These data suggest that H₂O₂ hastwo effects on the metabolism of flavonols: induction of theirsynthesis and stimulation of their oxidation. (Received March 6, 1989; Accepted July 10, 1989)     

Effect of volume expansion on hemodynamic variables in nephrectomized dogs

We have previously demonstrated that blood pressure elevation by acute blood volume expansion is volume-dependent during the infusion period and resistance-dependent in the post-infusion period in normal anesthetized dogs, and that such an increase in blood pressure is associated with a potentiation of the pressor response to norepinephrine. To evaluate the possible renal contribution to these hemodynamic changes, blood volume expansion was performed for 1 h with dextran dissolved in lactated Ringer's solution (20 ml/kg) in 15 nephrectomized dogs. The mean blood pressure, cardiac output and total peripheral resistance at the end of infusion were 126%, 225% and 60%, respectively; 3 h after volume expansion they were 126%, 151%, and 92% respectively. However, in 4 dogs, there was an increase in mean blood pressure (138%) 3 h after volume expansion. This was thought to result from an increase in the total peripheral resistance (133%) associated with the recovery of cardiac output (106%). The pressor response to norepinephrine (0.5 microgram/kg) was potentiated after volume expansion. These results indicate that the handling of volume by the kidney contributed to the maintenance of an elevated level of cardiac output. However, nephrectomy did not seem to interfere with the hemodynamic switching of the causative factor for blood pressure elevation from increased cardiac output to increased total peripheral resistance. Neither was the potentiation of pressor response to norepinephrine affected.     

Comparison of growth properties of carrot hairy root in various bioreactors

Summary Growth properties of carrot hairy root cells in various bioreactors were investigated. A turbine-blade reactor and an immobilized rotating drum reactor were found to be advantageous for the hairy root culture because of a high oxygen transfer coefficient (k in L a). After 30 days of culture, 10 g/l of dry hairy root cells were obtained in both bioreactors and maximum growth rates (V _m ) were found to be 0.63 and 0.61 g/l per day for the turbine-blade reactor and immobilized rotating drum reactor, respectively. Specific growth rates () at various cultivation times were observed to be linearly proportional to X/k _l a for both bioreactor configurations where X is the cell concentration. The estimated specific oxygen uptake rate of 0.34 mmol O₂/g dry cells per hour compares fairly well with an experimental value of 0.3.     

Prostaglandin D2 receptor of mastocytoma P-815 cells--possible regulation by phosphorylation and dephosphorylation

The 3H-labeled prostaglandin D2 [( 3H]PGD2) binding protein in the membrane fraction of mastocytoma P-815 cells was characterized. The specific binding of [3H]PGD2 to the cells or the membranes reached a maximum at pH 5.6, and was saturable, displaceable and of high affinity when incubated at 0 or 37 degrees C. The Bmax values for [3H]PGD2 binding in the two preparations at pH 5.6 were much higher at 0 degrees C than at 37 degrees C, whereas the Kd values were almost equal (85.3 nM for the cells and 80.5 nM for the membranes, respectively). High specific [3H]PGD2 binding activity in the mildly acid-treated cells was still observed when the external pH was raised from 5.6 to 7.2. Furthermore, specific [3H]PGD2 binding to the membranes (at 0 degrees C, pH 5.6) increased on addition of phosphatase inhibitors (NaF and molybdate) in the presence of 10 microM ATP, but practically disappeared on pretreatment of the membranes with phosphatase. On incubation of the membrane with [gamma-32P]ATP and molybdate, the stimulated incorporation of the [32P]phosphate into several peptides, including ones having an Mr of around 100,000-120,000, was observed. These results suggest that [3H]PGD2 binding in the mastocytoma P-815 cell membrane is controlled through phosphorylation-dephosphorylation of the receptor itself.     

Macrophage chemotactic factor (MCF) produced by a human T cell hybridoma clone

A human T cell hybridoma clone, D6-18, producing high levels of macrophage chemotactic factor (MCF) was established by the emetine-actinomycin D selection method. MCF was found to be present not only in the culture medium but also in the cell lysate of D6-18 cells. The secretion of the MCF from D6-18 cells was effectively inhibited by disodium cromoglycate, which is an inhibitor of the degranulation of mast cells, suggesting that MCF is stored in granules. The MCF of D6-18 cells was purified from the sonicated cell lysate by ion-exchange chromatographies and high-performance liquid chromatography. The amino acid sequence of the purified MCF was revealed to be WLGREDGSE or WLGRQDGSE. The synthetic peptide WLGREDGSE showed chemotactic activity against guinea pig macrophages and human monocytes at the concentration of about 10(-8) M.     

Mitochondrial DNA polymorphism in native Philippine cattle based on restriction endonuclease cleavage patterns

An analysis of patterns of cleavage of mtDNA by restriction endonucleases was performed for nine individuals from the Philippine population of native cattle. MtDNA polymorphisms were detected in the restriction patterns generated by the following six enzymes,BamHI,BglII,EcoRV,HindIII,PstI, andScaI. The restriction patterns showing polymorphisms were distributed nonrandomly among the nine individuals examined from the Philippine population of native cattle, indicating the existence of two separate types of mtDNA. These two types of mtDNA are very different from each other, at the level of subspecies. Since the native Philippine cattle are considered to represent an admixture of European and Indian cattle, the two types of mtDNA must be derived from the mtDNAs of both varieties. The polymorphic sites in mtDNA have been located on a restriction map, and the nucleotide substitutions at some of the sites have also been estimated.     

Molecular cloning of cDNA for proteasomes (multicatalytic proteinase complexes) from rat liver: primary structure of the largest component (C2)

Proteasomes (multicatalytic proteinase complexes) from rat liver are composed of at least 13 nonidentical components [Tanaka, K., Yoshimura, T., Ichihara, A., Ikai, A., Nishigai, M., Morimoto, M., Sato, M., Tanaka, N., Katsube, Y., Kameyama, K., & Takagi, T. (1988) J. Mol. Biol. 203, 985-996]. The nucleotide sequence of one major component (C2) of the proteasomes has been determined from a recombinant cDNA clone isolated by screening a rat liver cDNA library with a mixture of synthetic deoxyribonucleotides as a probe. The sequence was composed of 1174 nucleotides including a coding region for the entire protein and noncoding regions of both the 5'- and 3'-sides. The polypeptide deduced from the open reading frame consisted of 263 amino acid residues, and its molecular weight was calculated to be 29,516. The partial amino acid sequences of several fragments (approximately 45% of the total residues), which were obtained by cleavage of C2 with lysyl endopeptidase and cyanogen bromide, were determined by automated Edman degradation and found to be in complete accordance with those deduced from the cDNA sequence. The amino acid composition of C2, determined by chemical analysis, was also consistent with that deduced from the cDNA sequence, indicating that the cloned cDNA actually encoded component C2. Computer analysis revealed little structural similarity of C2 to other proteins reported so far. Northern blot hybridization analyses showed that the mRNA encoding this novel protein C2 was expressed in all the rat tissues examined and in a variety of eukaryotic organisms such as amphibia, birds, and mammals with slight species-specific differences in size.(ABSTRACT TRUNCATED AT 250 WORDS)     

Biosynthesis, processing and half-life of P-glycoprotein in a human multidrug-resistant KB cell

The biosynthesis, processing, and half-life of the drug efflux pump, P-glycoprotein, were studied in human multidrug-resistant KB (KB-C2) cells selected for resistance to colchicine. An antibody directed against a synthetic oligopeptide corresponding to the amino-acid sequence (Glu-393-Lys-408) of P-glycoprotein from human mdr1 cDNA was prepared in rabbits. With immunoblotting and immunoprecipitation, we detected a 140-170 kDa protein in KB-C2 cells but not in parental sensitive KB cells. KB-C2 cells made a 125 kDa precursor that was slowly processed (t1/2 = 45 min) to the mature form of 140-150 kDa. The processing rate of P-glycoprotein was slower than that of low-density lipoprotein receptor. We detected another 160-180 kDa smear band, which might be a completely denatured form of P-glycoprotein. With immunoblotting, a minor band of high molecular mass (greater than 500 kDa) was also detected and this form increased after the cells were treated with chemical cross-linker, 1,5-difluoro-2,4-dinitrobenzene. The half-life of P-glycoprotein was long; no significant loss of P-glycoprotein was observed within 24 h after synthesis. Cells treated with tunicamycin produced a 120 kDa form of P-glycoprotein which was no longer processed but showed stability similar to that of the mature 140-150 kDa form. Agents that reverse multidrug resistance, phorbol ester and transport substrate did not affect the stability of P-glycoprotein.     

Microfibrils: a constitutive component of reticular fibers in the mouse lymph node

Summary Fibrous components other than collagen fibrils in the reticular fiber of mouse lymph node were studied by electron microscopy. Bundles of microfibrils not associated by elastin and single microfibrils dispersed among collagen fibrils were present. The diameter of the microfibrils was 13.29±2.43 nm (n=100). Elastin-associated microfibrils occurred at the periphery of the reticular fiber. Elastin was enclosed by microfibrils, thus forming the elastic fiber, which was clearly demonstrated by tannic acid-uranyl acetate staining. In the reticular fiber of lymph nodes, the elastic fiber consisted of many more microfibrils and a small amount of elastin. These microfibrils, together with the collagen fibrils, may contribut to the various functions of the reticular fibers.