Mapping DNase-I hypersensitive sites on human isochores

Mapping DNase-I hypersensitive sites (HS) was used in the past to identify regulatory elements of specific genes. More recently, thousands of HS were identified in the human genome by using high-throughput methods. These approaches showed a general enrichment of HS near or within known genes, within CpG islands, within human-mouse conserved regions and in GC-rich regions of the genome. Here we show that HS: (i) are characterized by a much higher GC level (approximately 56%) than the average GC level of the human genome (approximately 41%); (ii) are overwhelmingly located in the GC-richest compartment of the genome, which is predominantly associated with an open chromatin structure; (iii) and are slightly more and slightly less frequent than genes, respectively, in the gene-rich and in the gene-poor isochore families.     

Is Cryopreservation a Homogeneous Process? Ultrastructure and Motility of Untreated, Prefreezing, and Postthawed Spermatozoa of Diplodus puntazzo (Cetti)

This study subdivides the cryopreservation procedure for Diplodus puntazzo spermatozoa into three key phases, fresh, prefreezing (samples equilibrated in cryosolutions), and postthawed stages, and examines the ultrastructural anomalies and motility profiles of spermatozoa in each stage, with different cryodiluents. Two simple cryosolutions were evaluated: 0.17 M sodium chloride containing a final concentration of 15% dimethyl sulfoxide (Me₂SO) (cryosolution A) and 0.1 M sodium citrate containing a final concentration of 10% Me₂SO (cryosolution B). Ultrastructural anomalies of the plasmatic and nuclear membranes of the sperm head were common and the severity of the cryoinjury differed significantly between the pre- and the postfreezing phases and between the two cryosolutions. In spermatozoa diluted with cryosolution A, during the prefreezing phase, the plasmalemma of 61% of the cells was absent or damaged compared with 24% in the fresh sample (P < 0.001). In spermatozoa diluted with cryosolution B, there was a pronounced increase in the number of cells lacking the head plasmatic membrane from the prefreezing to the postthawed stages (from 32 to 52%, P < 0.01). In both cryosolutions, damages to nuclear membrane were significantly higher after freezing (cryosolution A: 8 to 23%, P < 0.01; cryosolution B: 5 to 38%, P < 0.001). With cryosolution A, the after-activation motility profile confirmed a consistent drop from fresh at the prefreezing stage, whereas freezing and thawing did not affect the motility much further and 50% of the cells were immotile by 60–90 s after activation. With cryosolution B, only the postthawing stage showed a sharp drop of motility profile. This study suggests that the different phases of the cryoprocess should be investigated to better understand the process of sperm damage.     

Leaf morphology and macronutrients in broadleaved trees in central Italy

As part of an intensive monitoring programme (MON.I.TO, Intensive Monitoring of Forests in Tuscany), a 3-year survey was carried out, which included three tree species (beech, Fagus sylvatica L.; Turkey oak, Quercus cerris L.; holm-oak, Quercus ilex L.) located at six different sites. Leaves were sampled annually and analysed for nutrient concentrations (nitrogen, sulphur, phosphorus, magnesium, calcium, potassium and sodium) and morphological parameters (leaf area, dry weight, leaf mass per area, leaf thickness and leaf density). Results indicated considerable interannual variation of all the parameters. Differences between sampling sites indicated that, of all parameters measured, leaf mass per area could explain best the differences in field performance under conditions of stress. In fact, leaf mass per area was greater in the drier sites or when sea salt deposition occurred. Nevertheless, the variation of leaf mass per area over the 3 years did not reflect the differences in rainfall. Higher leaf mass per area was accompanied by lower concentrations of phosphorus and nitrogen, which could be a dilution effect due to an increase of structural carbon compounds in sclerophyllous leaves, as revealed by the total foliar content of these elements. Leaf mass per area as a measure of sclerophylly reached very high values among mesophile vegetation. Long-range transport of sea salt from coastal areas to mountain areas was mirrored in sodium concentrations of leaves.     

Identification of thieno[3,2-b]pyrroles as allosteric inhibitors of hepatitis C virus NS5B polymerase

Thieno[3,2-b]pyrroles are a novel class of allosteric inhibitors of HCV NS5B RNA-dependent RNA polymerase which show potent affinity for the NS5B enzyme. Introduction of a polar substituent in the position N1 led to a compound that efficiently blocks subgenomic HCV RNA replication in HUH-7 cells with an EC50 of 2.9 microM.     

Hyperglycemia in streptozotocin-induced diabetes leads to persistent inflammation and tissue damage following uveitis due to reduced levels of ciliary body heme oxygenase-1

This study investigated the heme oxygenase-1 (HO-1) and the endotoxin-induced uveitis (EIU) in diabetic streptozotocin (STZ)-hyperglycemic rats. STZ-hyperglycemic rats had impaired levels of the enzyme HO-1 within the ciliary bodies if compared with the nondiabetic rats. STZ-hyperglycemic rats also predisposed the eye to produce high levels of both the cytokines IL-1beta and CXCL8. Subsequent EIU further and significantly (P < .01) increased the cytokines production, an effect partly prevented by hemin treatment. Most importantly, hemin, an inducer of heme oxygenase expression and activity, recovered the huge number of infiltrated polymorphonuclear leukocytes PMN within the ciliary bodies associated with STZ-hyperglycemic state and EIU damage. Impairment of the stress-sensitive enzyme HO-1 in STZ-hyperglycemic rats increases and prolongs the inflammatory response to EIU.     

Chromosomal integration of LTR-flanked DNA in yeast expressing HIV-1 integrase: down regulation by RAD51

HIV-1 integrase (IN) is the key enzyme catalyzing the proviral DNA integration step. Although the enzyme catalyzes the integration step accurately in vitro, whether IN is sufficient for in vivo integration and how it interacts with the cellular machinery remains unclear. We set up a yeast cellular integration system where integrase was expressed as the sole HIV-1 protein and targeted the chromosomes. In this simple eukaryotic model, integrase is necessary and sufficient for the insertion of a DNA containing viral LTRs into the genome, thereby allowing the study of the isolated integration step independently of other viral mechanisms. Furthermore, the yeast system was used to identify cellular mechanisms involved in the integration step and allowed us to show the role of homologous recombination systems. We demonstrated physical interactions between HIV-1 IN and RAD51 protein and showed that HIV-1 integrase activity could be inhibited both in the cell and in vitro by RAD51 protein. Our data allowed the identification of RAD51 as a novel in vitro IN cofactor able to down regulate the activity of this retroviral enzyme, thereby acting as a potential cellular restriction factor to HIV infection.     

Skeletal muscle apoptosis is not increased in gastric cancer patients with mild-moderate weight loss

Numerous experimental and clinical studies have shown that skeletal muscle apoptotis may increase in wasting conditions and suggest that apoptosis might contribute to the loss of lean body mass. Data in cancer patients are still lacking. The present study aimed at verifying whether apoptosis was enhanced in the skeletal muscle of 16 patients with gastric cancer with respect to controls. A biopsy specimen was obtained from the rectus abdominis muscle. The occurrence of apoptosis in muscle biopsies was determined morphologically by the fluorescent transferase-mediated dUTP nick end labeling assay and by immunohistochemistry for caspase-3 and caspase-1. Mean weight loss was 6+/-2% in cancer patients and 0.5+/-0.1% in controls (p<0.0001). Serum albumin levels (g/dL) were 3.7+/-0.3 in cancer patients and 4.1+/-0.2 in controls (p<0.05). The percentage of apoptotic myonuclei was similar in cancer patients and in controls (1.5+/-0.3 versus 1.4+/-0.2, respectively; p=ns), in gastric cancer patients with mild (1.6+/-0.4) or moderate-severe weight loss (1.4+/-0.5) (p=ns), and in the different stages of disease (stages I-II: 1.5+/-0.7; stage III: 1.3+/-0.4; stage IV: 1.6+/-0.3; p=ns). By immunohistochemistry, caspase-1 and caspase-3 positive fibers were absent in controls and in neoplastic patients. Poly-ADP-ribosyl polymerase, a typical caspase-3 substrate whose processing is indicative of caspase-3 activation, was not cleaved in muscle biopsies of cancer patients. These data suggest that skeletal muscle apoptosis is not increased in neoplastic patients with mild-moderate weight loss and argue against the hypotheses that caspase-3 activation might be an essential step of myofibrillar proteolysis in cancer-related muscle wasting.     

The Function and Determinants of Reconciliation in <Emphasis Type="BoldItalic">Pan troglodytes</Emphasis>

Reconciliation (the postconflict affiliative reunion between former opponents) may mitigate costs of aggressive conflict by repairing the opponents’ relationship and reducing stress. We showed that postconflict levels of self-directed behavior were lower after reconciliation than when reconciliation did not occur in chimpanzees (Pan troglodytes) at Chester Zoo, providing support for a stress-alleviating function for reconciliation. Further, we investigated the effects of multiple factors on the occurrence of reconciliation using generalized linear mixed models. We performed 2 separate analyses, a “traditional” analysis and a “targeted” analysis. The former included variables previously used to assess the occurrence of reconciliation in primates, i.e., conflict characteristics, sex combination, and a simple measure of relationship value. The latter included species-specific variables such as the occurrence of consolation (postconflict affiliation from a bystander to the recipient of aggression); initiation of the conflict with a bluff display; and measures of relationship value, compatibility, and security specific to the study group. Whereas the traditional analysis showed that female-female dyads and valuable partners were most likely to reconcile, the targeted analysis showed that reconciliation was less likely to occur when consolation took place or when aggression was initiated with bluff displays. Further analyses revealed that the effect of sex-combination on reconciliation was due to its intercorrelation with bluff display. This study highlights the importance of considering variables specific to the study species and group when investigating the determinants of reconciliation and warns against premature interpretation of results without due consideration for all other possible determinants.     

Physiological adaptation in noncompetitive rock climbers: good for aerobic fitness?

The present investigation aimed to establish whether noncompetitive rock climbing fulfills sports medicine recommendations for maintaining a good level of aerobic fitness. The physiological profile of 13 rock climbers, 8 men (age, 43 +/- 8 years) and 5 women (age, 31 +/- 8 years) was assessed by means of laboratory tests. Maximal aerobic power (VO2peak) and ventilatory threshold (VT) were assessed using a cycloergometer incremental test. During outdoor rock face climbing, VO2 and heart rate (HR) were measured with a portable metabolimeter and the relative steady-state values (VO2 and HR during rock climbing) were computed. Blood lactate was measured during recovery. All data are presented as mean +/- SD. VO2 was 39.1 +/- 4.3 mL.kg.min in men and 39.7 +/- 5 mL.kg.min in women, while VT was 29.4 +/- 3.0 mL.kg.min in men and 28.8 +/- 4.6 mL.kg.min in women. The VO2 during rock climbing was 28.3 +/- 1.5 mL.kg.min in men and 27.5 +/- 3.7 mL.kg.min in women. The HR during rock climbing was 144 +/- 16 b.min in men and 164 +/- 13 b.min in women. The aerobic profile was classified from excellent to superior in accordance with the standards of the American College of Sports Medicine (ACSM). The exercise intensity (VO2 during rock climbing expressed as a percentage of VO2peak) was 70 +/- 6% in men and 72 +/- 8% in women. Moreover, the energy expenditure was 1000-1500 kcal per week. In conclusion, noncompetitive rock climbing has proved to be a typical aerobic activity. The intensity of exercise is comparable to that recommended by the American College of Sports Medicine to maintain good cardiorespiratory fitness.