A link between inflammation and metastasis: serum amyloid A1 and A3 induce metastasis, and are targets of metastasis-inducing S100A4.

S100A4 is implicated in metastasis and chronic inflammation, but its function remains uncertain. Here we establish an S100A4-dependent link between inflammation and metastatic tumor progression. We found that the acute-phase response proteins serum amyloid A (SAA) 1 and SAA3 are transcriptional targets of S100A4 via Toll-like receptor 4 (TLR4)/nuclear factor-κB signaling. SAA proteins stimulated the transcription of RANTES (regulated upon activation normal T-cell expressed and presumably secreted), G-CSF (granulocyte-colony-stimulating factor) and MMP2 (matrix metalloproteinase 2), MMP3, MMP9 and MMP13. We have also shown for the first time that SAA stimulate their own transcription as well as that of proinflammatory S100A8 and S100A9 proteins. Moreover, they strongly enhanced tumor cell adhesion to fibronectin, and stimulated migration and invasion of human and mouse tumor cells. Intravenously injected S100A4 protein induced expression of SAA proteins and cytokines in an organ-specific manner. In a breast cancer animal model, ectopic expression of SAA1 or SAA3 in tumor cells potently promoted widespread metastasis formation accompanied by a massive infiltration of immune cells. Furthermore, coordinate expression of S100A4 and SAA in tumor samples from colorectal carcinoma patients significantly correlated with reduced overall survival. These data show that SAA proteins are effectors for the metastasis-promoting functions of S100A4, and serve as a link between inflammation and tumor progression.

Perineal hernia repair using gracilis myocutaneous flap.

Perineal hernia formation is an infrequent but well-recognized complication of major pelvic surgery. Various methods of perineal reconstruction have been reported. This report describes one technique of perineal hernia repair using a unilateral gracilis myocutaneous flap. The gracilis myocutaneous flap provides well-vascularized tissue that is useful in many situations requiring reconstruction of the pelvis and perineum, especially when the area has been irradiated.

[Renal insufficiency in myelomatosis. Causes and treatment]. / Nyreinsufficiens ved myelomatose. Arsager og behandling.

Renal failure is a frequent complication in multiple myeloma and it is present in about 50% of patients with newly-diagnosed multiple myeloma. Renal failure at the time of diagnosis has earlier been associated with a bad prognosis, but a better prognostic factor is the response to chemotherapy. In general, it is important to distinguish between 1) renal insufficiency at the time of diagnosis, 2) acute renal insufficiency and 3) chronic renal insufficiency developing during the course of the disease. The patients in the first two groups are treated with intensive therapy which is long-lasting (median four to six weeks). The improved function of the kidney is correlated to an improved survival. Patients developing chronic renal insufficiency late in the course of the disease should receive palliative therapy. The most important factors that provoke acute renal insufficiency are dehydration, hypercalcaemia and/or infection, but renal insufficiency is also provoked by the use of nephrotoxic drugs, hyperuricaemia and/or hyperviscosity. Chronic renal insufficiency is provoked by deposits of light chains, infiltration by plasma cells or deposits of amyloid. The treatment consists of elimination of the provoking factors and start of chemotherapy.

A novel aspartyl protease allowing KEX2-independent MF alpha propheromone processing in yeast.

Mutants of Saccharomyces cerevisiae which lack the KEX2-encoded endopeptidase are unable to process proteolytically the mating factor alpha (MF alpha) propheromone produced from the chromosomal MF alpha 1 and MF alpha 2 genes (Julius et al., 1983). Overproduction of pheromone precursor from multiple, plasmid-borne MF alpha genes did, however, lead to the production of active MF alpha peptides in the absence of the KEX2 gene product. S. cerevisiae therefore must possess an alternative processing enzyme. The cleavage site of this enzyme appeared identical to that of the KEX2-encoded endopeptidase. To identify the gene responsible for the alternative processing, we have isolated clones which allowed production of mature MF alpha in a kex2-disrupted strain even from the chromosomal MF alpha genes. The gene isolated in this way was shown also to be essential for the KEX2-independent processing of propheromone overproduced from plasmid-borne MF alpha 1. The amino acid sequence deduced from the gene shows extensive homology to a number of aspartyl proteases including the PEP4 and BAR1 gene products from S. cerevisiae. In contrast to the BAR1 gene product, the novel aspartyl protease (YAP3 for Yeast Aspartyl Protease 3) contains a C-terminal serine/threonine-rich sequence and potential transmembrane domain similar to those found in the KEX2 gene product. The corresponding gene YAP3 was located to chromosome XII. The normal physiological role of the YAP3 gene product is not known. Strains disrupted in YAP3 are both viable and able to process the mating factor a precursor.

The secretion of glucagon by transformed yeast strains.

Saccharomyces cerevisiae strains were transformed with plasmids coding for modified mating factor alpha 1 leader sequences followed by glucagon. Glucagon-containing peptides which were secreted into the fermentation broth were isolated and their amino acid sequences determined. The yeast strain transformed with the sequence coding for the complete mating factor alpha 1 leader sequence preceding the glucagon gene (MT556) secreted glucagon plus glucagon extended at its N-terminal by parts of the leader sequence. The yeast strain transformed with the sequence coding for a truncated mating factor alpha 1 leader sequence before the glucagon gene (MT615) secreted glucagon. These observations suggest that S. cerevisiae is a suitable vehicle for the efficient expression of plasmids coding for polypeptides similar to glucagon (e.g. VIP, secretin, GIP).

Sensitivity of Escherichia coli acrA mutants to psoralen plus near-ultraviolet radiation.

The sensitivity to psoralen plus near-ultraviolet radiation (PUVA) was compared in a pair of E. coli strains differing at the acrA locus. Survival was determined for both bacteria and phage lambda. AcrA mutant cells were 40 times more sensitive than wild type to the lethal effect of PUVA. Free lambda phage exposed to PUVA survived as well when plated on acrA mutants as on wild type. In contrast, prophage lambda CI857 ind carried in lysogenic acrA strains was hypersensitive to PUVA. The enhanced sensitivity of bacterial and lambda DNA, when inside acrA cells, was paralleled by an increased photobinding of radiolabelled psoralens in the mutant. Binding was increased specifically to DNA rather than to nucleic acids in general. The difference in psoralen-binding ability determined by the acrA gene persisted after permeabilizing treatment of the cells. The results suggest that the acrA mutation causes an alteration specifically in the environment of the cellular DNA so as to allow increased intercalation and photobinding of psoralens.

Rescue of mitomycin C- or psoralen-inactivated Micrococcus radiodurans by additional exposure to radiation or alkylating agents.

The processing of damaged DNA was altered in a mitomycin C-sensitive mutant (mtcA) of Micrococcus radiodurans. Even though the mutant retained resistance to 254-nm UV radiation, it did not, in contrast to the wild-type strain, show any excessive DNA degradation or cell death when incubated with chloramphenicol after sublethal doses of either UV light or mitomycin C. The results suggest the constitutive synthesis of an enzyme system responsible for wild-type proficiency in the repair of mitomycin C-induced damage. An alternative system able to repair damage caused by mitomycin C was demonstrated in the mtcA background. In this strain, additional damage inflicted upon the cellular DNA effected a massive rescue of cells previously inactivated by mitomycin C. Rescue was provoked by ionizing radiation, by UV light, or by simple alkylating agents. Cells treated with psoralen plus near-UV radiation could be rescued only when inactivation was due primarily to psoralen-DNA interstrand cross-links rather than to monoadducts. The rescue of inactivated cells was prevented in the presence of chloramphenicol. These results can be interpreted most readily in terms of an alternative repair system able to overcome DNA interstrand cross-links produced by mitomycin C or psoralen plus near-UV light, but induced only by the more abundant number of damages produced by radiation or simple alkylating agents.

Multiplicity of genome equivalents in the radiation-resistant bacterium Micrococcus radiodurans.

The complexity of the genome of Micrococcus radiodurans was determined to be (2.0 +/- 0.3) X 10(9) daltons by DNA renaturation kinetics. The number of genome equivalents of DNA per cell was calculated from the complexity and the content of DNA. A lower limit of four genome equivalents per cell was approached with decreasing growth rate. Thus, no haploid stage appeared to be realized in this organism. The replication time was estimated from the kinetics and amount of residual DNA synthesis after inhibiting initiation of new rounds of replication. From this, the redundancy of terminal genetic markers was calculated to vary with growth rate from four to approximately eight copies per cell. All genetic material, including the least abundant, is thus multiply represented in each cell. The potential significance of the maintenance in each cell of multiple gene copies is discussed in relation to the extreme radiation resistance of M. radiodurans.

Simple downshift and resulting lack of correlation between ppGpp pool size and ribonucleic acid accumulation.

The growth rate of Escherichia coli can be limited by the availability of carbon and energy. To impose such a limitation, alpha-methylglucoside (alpha MG), a non-metabolizable analogue, can be used to decrease uptake of glucose by competition for the transport of this sugar. Varying the ratio of glucose to alphaMG allowed shifts in growth rate without simultaneous qualitative changes in the growth medium and permitted examination of the immediate changes accompanying such shifts. Stringent (rel+) as well as relaxed (rel minus) strains were able to rapidly curtail their accumulation of ribonculeic acid (RNA) after a downshift imposed by decreasing glucose transport into the cell. Guanosine 5'-diphosphate 3'-diphosphate (ppGpp) accumulated in both rel+ and rel minus strains after a degrease in growth rate. However, the accumulation of ppGpp in relaxed derivatives was very slow, and there was no direct or obligatory correlation between the level of ppGpp and the rate of RNA accumulation. This latter conclusion is supported by measurements of ppGpp levels and rates of RNA accumulation after restoration of maximal growth rates by addition of excess glucose.

Influence of polyamine limitation on the chain growth rates of beta-galactosidase and of its messenger ribonucleic acid.

The rates of elongation of beta-galactosidase and its messenger ribonucleic acid (RNA) were estimated in a polyamine-deficient mutant of Escherichia coli through an analysis of the kinetics of enzyme induction. The chain growth of beta-galactosidase was calculated from the time required after the appearance of an amino terminal fragment of 60 amino acids (auto-alpha) until completed enzyme began to accumulate. The elongation rate of beta-galactosidase messenger RNA was estimated from the time after induction at which streptolydigen-resistant, enzyme-forming capacity first appeared. Upon polyamine starvation, the rate of polypeptide elongation slowed from 17 to 10 amino acids per s and the messenger RNA elongation rate decreased from 47 to 30 nucleotides per s. These reductions in polymerization rates were proportional to the decrease in cellular growth rate produced by polyamine starvation. It was concluded that, although it is quite unlikely that polyamine levels are involved in regulation of cell growth, they may be acting as cofactors in the synthesis of RNA or protein, or both.