Acute physical exercise and long-term individual shear rate therapy increase telomerase activity in human peripheral blood mononuclear cells.

AIM: Physical activity is a potent way to impede vascular ageing. However, patients who suffer from peripheral artery disease (PAD) are often unable to exercise adequately. For those patients, we have developed individual shear rate therapy (ISRT), which is an adaptation of external counterpulsation and enhances endovascular fluid shear stress to increase collateral growth (arteriogenesis). To evaluate the effects of physical exercise and ISRT on the telomere biology of peripheral blood mononuclear cells (PBMCs), we conducted two clinical trials. METHODS: In the ISRT-1 study, we assessed PBMC telomerase activity in 26 young healthy volunteers upon a single (short-term) ISRT session and a single treadmill running session. In the ISRT-2 study, we investigated PBMC telomere biology of 14 elderly patients with PAD, who underwent 30 h of (long-term) ISRT within a 5-week period. RESULTS: We demonstrate that telomerase activity significantly increased from 39.84 Total Product Generated (TPG) Units ± 6.15 to 58.10 TPG ± 10.46 upon a single treadmill running session in healthy volunteers. In the ISRT-2 trial, PBMC telomerase activity and the mRNA expression of the telomere-protective factor TRF2 increased from 40.87 TPG ± 4.45 to 60.98 TPG ± 6.83 and 2.10-fold ± 0.40, respectively, upon long-term ISRT in elderly patients with PAD. CONCLUSION: In summary, we show that acute exercise and long-term ISRT positively affect PBMC telomerase activity, which is indicative for an improved regenerative potential of immune cells and vascular tissues. Long-term ISRT also enhances the gene expression of the telomere-protective factor TRF2.

PPARγ activation inhibits cerebral arteriogenesis in the hypoperfused rat brain.

AIMS: PPARγ stimulation improves cardiovascular (CV) risk factors, but without improving overall clinical outcomes. PPARγ agonists interfere with endothelial cell (EC), monocyte and smooth muscle cell (SMC) activation, function and proliferation, physiological processes critical for arterial collateral growth (arteriogenesis). We therefore assessed the effect of PPARγ stimulation on cerebral adaptive and therapeutic collateral growth. METHODS: In a rat model of adaptive cerebral arteriogenesis (3-VO), collateral growth and function were assessed (i) in controls, (ii) after PPARγ stimulation (pioglitazone 2.8 mg kg(-1); 10 mg kg(-1) compared with metformin 62.2 mg kg(-1) or sitagliptin 6.34 mg kg(-1)) for 21 days or (iii) after adding pioglitazone to G-CSF (40 µg kg(-1) every other day) to induce therapeutic arteriogenesis for 1 week. Pioglitazone effects on endothelial and SMC morphology and proliferation, monocyte activation and migration were studied. RESULTS: PPARγ stimulation decreased cerebrovascular collateral growth and recovery of hemodynamic reserve capacity (CVRC controls: 12 ± 7%; pio low: -2 ± 9%; pio high: 1 ± 7%; metformin: 9 ± 13%; sitagliptin: 11 ± 12%), counteracted G-CSF-induced therapeutic arteriogenesis and interfered with EC activation, SMC proliferation, monocyte activation and migration. CONCLUSION: Pharmacologic PPARγ stimulation inhibits pro-arteriogenic EC activation, monocyte function, SMC proliferation and thus adaptive as well as G-CSF-induced cerebral arteriogenesis. Further studies should evaluate whether this effect may underlie the CV risk associated with thiazolidinedione use in patients.

Lethal mutations defining 112 complementation groups in a 4.5 Mb sequenced region of Caenorhabditis elegans chromosome III.

The central gene cluster of chromosome III was one of the first regions to be sequenced by the Caenorhabditis elegans genome project. We have performed an essential gene analysis on the left part of this cluster, in the region around dpy-17III balanced by the duplication sDp3. We isolated 151 essential gene mutations and characterized them with regard to their arrest stages. To facilitate positioning of these mutations, we generated six new deficiencies that, together with preexisting chromosomal rearrangements, subdivide the region into 14 zones. The 151 mutations were mapped into these zones. They define 112 genes, of which 110 were previously unidentified. Thirteen of the zones have been anchored to the physical sequence by polymerase chain reaction deficiency mapping. Of the 112 essential genes mapped, 105 are within these 13 zones. They span 4.2 Mb of nucleotide sequence. From the nucleotide sequence data, 920 genes are predicted. From a Poisson distribution of our mutations, we predict that 234 of the genes will be essential genes. Thus, the 105 genes constitute 45% of the estimated number of essential genes in the physically defined zones and between 2 and 5% of all essential genes in C. elegans.

Interpreting a sequenced genome: toward a cosmid transgenic library of Caenorhabditis elegans.

We have generated a library of transgenic Caenorhabditis elegans strains that carry sequenced cosmids from the genome of the nematode. Each strain carries an extrachromosomal array containing a single cosmid, sequenced by the C. elegans Genome Sequencing Consortium, and a dominate Rol-6 marker. More than 500 transgenic strains representing 250 cosmids have been constructed. Collectively, these strains contain approximately 8 Mb of sequence data, or approximately 8% of the C. elegans genome. The transgenic strains are being used to rescue mutant phenotypes, resulting in a high-resolution map alignment of the genetic, physical, and DNA sequence maps of the nematode. We have chosen the region of chromosome III deleted by sDf127 and not covered by the duplication sDp8(III;I) as a starting point for a systematic correlation of mutant phenotypes with nucleotide sequence. In this defined region, we have identified 10 new essential genes whose mutant phenotypes range from developmental arrest at early larva, to maternal effect lethal. To date, 8 of these 10 essential genes have been rescued. In this region, these rescues represent approximately 10% of the genes predicted by GENEFINDER and considerably enhance the map alignment. Furthermore, this alignment facilitates future efforts to physically position and clone other genes in the region. [Updated information about the Transgenic Library is available via the Internet at http://darwin.mbb.sfu.ca/imbb/dbaillie/cos mid.html.]

The circum-Chryse region as a possible example of a hydrologic cycle on Mars: geologic observations and theoretical evaluation.

The transection and superposition relationships among channels, chaos, surface materials units, and other features in the circum-Chryse region of Mars were used to evaluate relative age relationships and evolution of flood events. Channels and chaos in contact (with one another) were treated as single discrete flood-carved systems. Some outflow channel systems form networks and are inferred to have been created by multiple flood events. Within some outflow channel networks, several separate individual channel systems can be traced to a specific chaos which acted as flood-source area to that specific flood channel. Individual flood-carved systems were related to widespread materials units or other surface features that served as stratigraphic horizons. Chryse outflow channels are inferred to have formed over most of the perceivable history of Mars. Outflow channels are inferred to become younger with increasing proximity to the Chryse basin. In addition, outflow channels closer to the basin show a greater diversity in age. The relationship of subsequent outflow channel sources to the sources of earlier floods is inferred to disfavor episodic flooding due to the progressive tapping of a juvenile near-surface water supply. Instead, we propose the circum-Chryse region as a candidate site of past hydrological recycling. The discharge rates necessary to carve the circum-Chryse outflow channels would have inevitably formed temporary standing bodies of H2O on the Martian surface where the flood-waters stagnated and pooled (the Chryse basin is topographically enclosed). These observations and inferences have led us to formulate and evaluate two hypotheses: (1) large amounts of the sublimated H2O off the Chryse basin flood lakes precipitated (snowed) onto the flood-source highlands and this H2O was incorporated into the near surface, recharging the H2O sources, making possible subsequent deluges; and (2) ponded flood-water in Chryse basin drained back down an anti basinward dipping subsurface layer accessed long the southern edge of the lake, recharging the flood-source aquifers. H2O not redeposited in the flood-source region was largely lost to the hydrologic cycle. This loss progressively lowered the vitality of the cycle, probably by now killing it. Our numerical evaluations indicate that of the two hypotheses we formulated, the groundwater seep cycle seems by far the more viable. Optimally, approximately 3/4 of the original mass of an ice-covered cylindrical lake (albedo 0.5, 1 km deep, 100-km radius, draining along its rim for one quarter of its circumference into substrata with a permeability of 3000 darcies) can be modeled to have moved underground (on timescales of the order of 10(3) years) before the competing mechanisms of sublimation and freeze down choked off further water removal. Once underground, this water can travel distances equal to the separation between Chryse basin and flood-source sites in geologically short (approximately 10(6) year-scale) times. Conversely, we calculate that optimally only approximately 40% of the H2O carried from Chryse can condense at the highlands, and most of the precipitate would either collect at the base of the highlands/lowlands scarp or sublimate at rates greater than it would accumulate over the flood-source sites. Further observations from forthcoming missions may permit the determination of which mechanisms may have operated to recycle the Chryse flood-waters.

Hydroxylation and Dechlorination of Tetrachlorohydroquinone by Rhodococcus sp. Strain CP-2 Cell Extracts.

A cell extract of a polychlorophenol-degrading bacterium, Rhodococcus sp. strain CP-2, isolated from chlorophenol-contaminated soil, was shown to dechlorinate tetrachlorohydroquinone, the first intermediate in pentachlorophenol and 2,3,5,6-tetrachlorophenol degradation. Degradation of tetrachlorohydroquinone was catalyzed by a soluble enzyme(s). The reaction sequence for complete dechlorination involved hydroxylation and three reductive dechlorinations, producing 1,2,4-trihydroxybenzene. All chlorines were thus removed from the polychlorinated compound before ring cleavage.

Transformation of chlorinated phenolic compounds in the genusRhodococcus.

The ability of strains of the genusRhodococcus to transform chlorinated phenolic compounds was studied. Noninduced cells of several strains ofRhodococcus, covering at least eight species, were found to attack mono-, di-, and trichlorophenols by hydroxylation at theortho position to chlorocatechols. 3-chlorophenol and 4-chlorophenol were converted to 4-chlorocatechol, 2,3-dichlorophenol to 3,4-dichlorocatechol, and 3,4-di-chlorophenol to 4,5-dichlorocatechol. The chlorocatechols accumulated to nearly stoichiometric amounts. Other mono- and dichlorophenols were not transformed. The ability of the strains to hydroxylate chlorophenols correlated with the ability to grow on unsubstituted phenol as the sole source of carbon and energy. SeveralRhodococcus strains attacked chlorophenolic compounds by both hydroxylation and O-methylation. 2,3,4-, 2,3,5- and 3,4,5-trichlorophenol were hydroxylated to trichlorocatechol and then sequentially O-methylated to chloroguaiacol and chloroveratrole. Tetrachlo-rohydroquinone was O-methylated sequentially to tetrachloro-4-methoxy-phenol and tetrachloro-1,4-dimethoxybenzene. Several of the active strains had no known history of exposure to any chloroaromatic compound. Rhodococci are widely distributed in soil and sludge and these results suggest that this genus may play an important role in transformation of chlorinated phenolic compounds in the environment.

In vivo generation of R68.45-pPGH1 hybrid plasmids conferring a Phl+ (meta pathway) phenotype.

Plasmid pPGH1 originating from Pseudomonas putida strain H carries all the genes required for the degradation of phenol (or cresols) via the meta cleavage pathway. Besides mobilization of pPGH1 by a plasmid of the incompatibility group P-1, hybrid plasmids conferring the Phl+ phenotype could be selected, when R68.45 was the conjugative plasmid. The hybrids contain the complete R68.45 and part of pPGH1. Integration of Phl-DNA of pPGH1 into R68.45 occurred exclusively via the IS21 region of R68.45.

Use of salicylate to estimate the "threshold" inducer level for de novo synthesis of the phenol-degrading enzymes in Pseudomonas putida strain H.

A special approach was used to elucidate the "threshold" inducer concentration for coordinative de novo synthesis of phenol hydroxylase(s), catechol 2,3-dioxygenase and the 2-hydroxymuconic semialdehyde-metabolizing enzymes which initiate phenol catabolism in Pseudomonas putida strain H. It is based on cell-precultivation with glucose (as the carbon and energy source) in the presence of different concentrations of sodium salicylate which proved to be a potent non-metabolizable inducer in strain H of these enzymes. Subsequent estimation of the activity status of resting cell suspensions and cell-free extracts, respectively, prepared from those strain H cultures clearly revealed failing de novo synthesis of the mentioned phenol-degrading enzymes at salicylate concentrations lower than 0.2 mg/l.

Nature and significance of microbial cometabolism of xenobiotics.

Microbial cometabolism, i.e. "transformation of a non-growth substrate in the obligate presence of a growth substrate or another transformable compound" (Dalton and Stirling 1982) is a whole-cell phenomenon physiologically based on coupling of different catabolic pathways at the cellular level. It is frequently observed in transformation of xenobiotic non-growth substrates by individual microbial species. Transformation processes of this type are usually mediated by appropriate non-specific enzymes of the peripheric cellular metabolism able to modify a variety of substances other than their natural substrates. The precise mechanisms of coupling between metabolism of xenobiotic non-growth substrates and of particular additional carbon substrates may be different depending on the substrates and the microbial species involved. However, experimental data indicate that the primary function of the respective additional carbon substrates is to supply either energy, cofactors or metabolites for the different cellular events involved in the transformation process (e.g. uptake of the xenobiotic non-growth substrate, functioning of appropriate degradative enzymes of the peripheric cellular metabolism). Cometabolism of xenobiotics involves nothing special or novel from the standpoint of biochemistry. On the contrary, there are numerous examples where the turnover of particular natural compounds by certain aerobic or anaerobic microorganisms is essentially based on coupling of different catabolic pathways at the cellular level by transfer of hydrogen (i.e. reducing power) and/or energy between two or more enzymatic reactions. Synthetic chemicals which resist total degradation by individual microbial species may undergo mineralization due to complementary catabolic sequences mediated by certain multispecies microbial associations with cometabolic transformations being the initial steps. Although taking place in certain natural habitats (e.g. rhizospheres, sewage), microbial cometabolism of xenobiotics in natural ecosystems occurs with slow rates since the respective cometabolizing populations are generally small and will not increase in number or biomass in response to the introduced chemicals. However, under conditions of axenic microbial cultures, high concentrations of biomass, and appropriate substrate mixtures cometabolism of synthetic chemicals may be a useful technique of considerable practical importance to accumulate biochemical products at high yields. In addition, cometabolic capabilities of wild-type microorganisms may serve as a tool for the construction of microbial strains with a new degradative potential for recalcitrant xenobiotic compounds.

Degradation of aniline and monochloroanilines by Rhodococcus sp. An 117 and a pseudomonad: a comparative study.

Two newly isolated aniline-degrading bacterial strains were characterized with regard to their enzyme systems responsible for aniline catabolism. One of them identified as a Rhodococcus sp. metabolized aniline exclusively via the beta-ketoadipate pathway by means of inducible enzymes. The aniline-degrading enzyme system of the second isolate, presumably a pseudomonad, was shown to consist of an inducible aniline-converting enzyme and constitutive meta-pathway enzymes. Both isolates failed to metabolize monochlorinated anilines in the absence of additional carbon sources. To explain this the ring-cleaving enzymes of both isolates were examined for their substrate specificities. Furthermore, the effect of 4-chlorocatechol on the enzymes catalyzing aniline conversion and catechol oxygenation was investigated.

Chromosome anomalies in 136 couples with a history of recurrent abortions.

Cytogenetic studies were performed on 136 couples with a history of two or more abortions referred to us after gynaecological causes of the abortions had been excluded. Fifteen (11%) of the couples were found to have a chromosome anomaly, and when the couples were subdivided according to number of abortions, surprisingly 6 (10%) of the 59 couples with a history of only two abortions had a chromosome anomaly. An increased frequency of mosaicism for X-chromosome aneuploidy (2.2%) in the women from the 136 couples was also found. A review of the literature shows that translocations of some chromosomes (e.g. nos. 1, 7 or 22) preferentially lead to fetal wastage, while those involving, for example, chromosome nos. 5, 9, 14 or 21 are more likely to result in the birth of a handicapped child. Couples with a history of two abortions should be investigated cytogenetically. Other causes of miscarriages must, however, be excluded first.

Regulation of phenol degradation in Pseudomonas putida.

In order to characterize the ability of Pseudomonas putida (TREVISAN 1889) MIGULA 1895 strain H to degrade various mono- and diphenolic aromatic compounds, respiratory activities towards phenol, catechol, and the cresol isomers were determined. The following rates of oxygen uptake (QO2) were obtained with resting phenol-grown cells: phenol -- 229, o-cresol -- 231, m-cresol -- 43, p-cresol -- 200, catechol -- 262. All these compounds were oxidized by a two-phase-kinetics, the first phase is characterized by a higher oxidation rate than the second. The oxidation of phenol as well as of p-cresol was found to be substrate-inhibited at concentrations above 0.25 mM. A Ki-value of 100 mg/l was calculated for phenol oxidation. The phenol-degrading enzyme system is induced, probably coordinately, by phenol and the cresol isomers. In strain H the degradation of phenol is carried out simultaneously with the assimilation of natural carbohydrates like glucose and sodium pyruvate. Aniline as well as sodium benzoate, though not metabolized by strain H, cause a concentration-dependent inhibition of phenol degradation in resting phenol-grown cells of that strain. The mechanism of this inhibition is discussed.

[District physician and family planning]. / Bereichsarzt und Familienplanung

PIP: The marriage and family planning counseling duties of the Bereichsarzt (area physician) in the German Democratic Republic are described. He works closely with the marriage and family counseling services of his area as a medical specialist concerned with the physiological and psychological aspects of marriage and family life. He refers patients to the appropriate specialized agencies as a part of his general duties in preventive medicine. In addition, the area physician helps to train counseling personnel in sexual physiology and psychology, equip the counseling center, and coordinate the work of several specialists on a single case.^ieng