Pulmonary diffusing capacity in reptiles (Relations to temperature and O2-uptake)

Summary Pulmonary CO-diffusing capacity (D l _CO), lung volume, pulmonary perfusion and O₂-uptake were measured by non-invasive techniques in the lizardsVaranus exanthematicus andTupinambis teguixin (mean body weight 2.2 kg for both species).The CO-diffusing capacity was at 25–27°C 0.059 mlstpd·kg^–1·min^–1·Torr^–1 inVaranus, which is 47% greater than the value of 0.040 mlstpd·kg^–1·min^–1·Torr^–1 inTupinambis. The lung volume ofVaranus was 36 ml·kg^–1 and that ofTupinambis 20 ml·kg^–1. At 35–37°C the diffusing capacity of lizard lungs are about 25% of those for mammals of comparable size.InVaranus pulmonary CO-diffusing capacity increased with temperature from 0.027 mlstpd·kg^–1·min^–1·Torr^–1 at 17–19 °C to 0.075 mlstpd·kg^–1·min^–1·Torr^–1 at 35–37 °C. This change closely matched a concomitant increase of O₂-uptake. Pulmonary perfusion increased from 27 ml·kg^–1·min^–1 to 55 ml·kg^–1·min^–1 within this temperature range.The study emphasizes that pulmonary diffusing capacity cannot be fully evaluated without information on pulmonary perfusion and O₂-uptake. In reptiles and other ectotherms diffusing capacity must be reported at specified body temperature.     

Antirotavirus Immunoglobulin A Neutralizes Virus In Vitro after Transcytosis through Epithelial Cells and Protects Infant Mice from Diarrhea

Rotaviruses are the major cause of severe diarrhea in infants and young children worldwide. Due to their restricted site of replication, i.e., mature enterocytes, local intestinal antibodies have been proposed to play a major role in protective immunity. Whether secretory immunoglobulin A (IgA) antibodies alone can provide protection against rotavirus diarrhea has not been fully established. To address this question, a library of IgA monoclonal antibodies (MAbs) previously developed against different proteins of rhesus rotavirus was used. A murine hybridoma “backpack tumor” model was established to examine if a single MAb secreted onto mucosal surfaces via the normal epithelial transport pathway was capable of protecting mice against diarrhea upon oral challenge with rotavirus. Of several IgA and IgG MAbs directed against VP8 and VP6 of rotavirus, only IgA VP8 MAbs (four of four) were found to protect newborn mice from diarrhea. An IgG MAb recognizing the same epitope as one of the IgA MAbs tested failed to protect mice from diarrhea. We also investigated if antibodies could be transcytosed in a biologically active form from the basolateral domain to the apical domain through filter-grown Madin-Darby canine kidney (MDCK) cells expressing the polymeric immunoglobulin receptor. Only IgA antibodies with VP8 specificity (four of four) neutralized apically administered virus. The results support the hypothesis that secretory IgA antibodies play a major role in preventing rotavirus diarrhea. Furthermore, the results show that the in vivo and in vitro methods described are useful tools for exploring the mechanisms of viral mucosal immunity.     

A comparison of risk factors for cryptosporidiosis and non-cryptosporidiosis diarrhoea: A case-case-control study in Ethiopian children

BackgroundCryptosporidiosis is a major cause of diarrhoea in young children in low-and-middle-income countries. New interventions should be informed by evidence pertaining to risk factors and their relative importance. Inconsistencies in the literature may to some extent be explained by choice of methodology, furthermore, most previous risk factor studies compared cryptosporidiosis cases to diarrhoea cases of other aetiologies rather than with controls without diarrhoea.Methodology/Principal findingsWe investigated a broad set of factors in under-2-year-olds presenting with diarrhoea to a hospital and a health center in southwestern Ethiopia. We applied quantitative cut-offs to distinguish between cryptosporidiosis and incidental Cryptosporidium infection or carriage, a hierarchical causal framework to minimize confounding and overadjustment, and a case-case-control design, to describe risk factors for both cryptosporidiosis and non-cryptosporidiosis diarrhoea. Moderate and severe acute malnutrition were strongly associated with both cryptosporidiosis and non-cryptosporidiosis diarrhoea. Previous healthcare attendance and low maternal education were only associated with cryptosporidiosis, whereas unsafe child stool disposal, prematurity and early cessation of exclusive breastfeeding were significantly associated with non-cryptosporidiosis diarrhoea only. By estimation of population attributable fractions, socioeconomic factors—specifically low maternal education—and public tap water use, were apparently more important risk factors for cryptosporidiosis than for non-cryptosporidiosis diarrhoea.Conclusions/SignificanceNutritional management of moderate acute malnutrition may be an effective intervention against cryptosporidiosis, particularly if combined with targeted therapy for cryptosporidiosis which, again, may mitigate nutritional insult. Focused caregiver education in healthcare settings and follow-up of children with acute malnutrition may prevent or improve outcomes of future episodes of cryptosporidiosis.     

The Genomic HyperBrowser: inferential genomics at the sequence level

The immense increase in the generation of genomic scale data poses an unmet analytical challenge, due to a lack of established methodology with the required flexibility and power. We propose a first principled approach to statistical analysis of sequence-level genomic information. We provide a growing collection of generic biological investigations that query pairwise relations between tracks, represented as mathematical objects, along the genome. The Genomic HyperBrowser implements the approach and is available at http://hyperbrowser.uio.no.     

Body composition and breast cancer in postmenopausal women: a Danish prospective cohort study

Objective: To assess the importance of body fat mass (BFM) and fat free mass (FFM) for the established positive association between BMI and breast cancer among post‐menopausal women. Research Methods and Procedures: A prospective cohort of 23,788 postmenopausal women included in the Danish study Diet, Cancer, and Health during 1993 to 1997 was linked to the Danish Cancer Registry to identify all cases of breast cancer occurring during 1993 to 2002. Breast cancer incidence rate ratios for anthropometric measurements with adjustment for known risk factors for breast cancer were calculated by Cox regression analyses. Results: Among the most commonly used anthropometric measurements, BMI was positively associated with breast cancer among never users of hormone replacement therapy (HRT). By splitting BMI into two indices, BFM index and FFM index, we found that the incidence rate ratio with each 1 kg/m² among never users of HRT was 0.98 (95% confidence interval, 0.93 to 1.03) for BFM index and 1.12 (95% confidence interval, 1.00 to 1.26) for FFM index after mutual adjustment. Discussion: The finding for BMI was in accordance with previous findings. Our results indicate that the FFM component of BMI may play a role for development of breast cancer among never users of HRT.     

The ins and outs of plant cell walls

New findings reveal that many membrane proteins undergo regulated trafficking between intracellular compartments and the plasma membrane. This also appears to be a common regulatory mechanism in the control of cell wall metabolism.     

Does multiple hosts mean multiple parasites? Population genetic structure of Schistosoma japonicum between definitive host species

Multi-host parasites, those capable of infecting more than one species of host, are responsible for the majority of all zoonotic, emerging or persistent human and animal diseases and are considered one of the major challenges for the biomedical sciences in the 21st century. We characterized the population structure of the multi-host parasite Schistosoma japonicum in relation to its definitive host species by genotyping miracidia collected from humans and domestic animals across five villages around the Yangtze River in Anhui Province, mainland China, using microsatellite markers. High levels of polymorphisms were observed and two main genetic clusters were identified which separated water buffalo, cattle and humans from goats, pigs, dogs and cats. We thereby believe that we present the first evidence of definitive host-based genetic variation in Schistosoma japonicum which has important epidemiological, evolutionary, medical and veterinary implications.     

Aurothiomalate inhibits transformed growth by targeting the PB1 domain of protein kinase Ciota

We recently identified the gold compound aurothiomalate (ATM) as a potent inhibitor of the Phox and Bem1p (PB1)-PB1 domain interaction between protein kinase C (PKC) iota and the adaptor molecule Par6. ATM also blocks oncogenic PKCiota signaling and the transformed growth of human lung cancer cells. Here we demonstrate that ATM is a highly selective inhibitor of PB1-PB1 domain interactions between PKCiota and the two adaptors Par6 and p62. ATM has no appreciable inhibitory effect on other PB1-PB1 domain interactions, including p62-p62, p62-NBR1, and MEKK3-MEK5 interactions. ATM can form thio-gold adducts with cysteine residues on target proteins. Interestingly, PKCiota (and PKCzeta) contains a unique cysteine residue, Cys-69, within its PB1 domain that is not present in other PB1 domain containing proteins. Cys-69 resides within the OPR, PC, and AID motif of PKCiota at the binding interface between PKCiota and Par6 where it interacts with Arg-28 on Par6. Molecular modeling predicts formation of a cysteinyl-aurothiomalate adduct at Cys-69 that protrudes into the binding cleft normally occupied by Par6, providing a plausible structural explanation for ATM inhibition. Mutation of Cys-69 of PKCiota to isoleucine or valine, residues frequently found at this position in other PB1 domains, has little or no effect on the affinity of PKCiota for Par6 but confers resistance to ATM-mediated inhibition of Par6 binding. Expression of the PKCiota C69I mutant in human non-small cell lung cancer cells confers resistance to the inhibitory effects of ATM on transformed growth. We conclude that ATM inhibits cellular transformation by selectively targeting Cys-69 within the PB1 domain of PKCiota.     

Phosphorylation of Thr654 but not Thr669 within the juxtamembrane domain of the EGF receptor inhibits calmodulin binding

Calcium-calmodulin (CaM) binding to the epidermal growth factor receptor (EGFR) has been shown to both inhibit and stimulate receptor activity. CaM binds to the intracellular juxtamembrane (JM) domain (Met645-Phe688) of EGFR. Protein kinase C (PKC) mediated phosphorylation of Thr654 occurs within this domain. CaM binding to the JM domain inhibits PKC phosphorylation and conversely PKC mediated phosphorylation of Thr654 or Glu substitution of Thr654 inhibits CaM binding. A second threonine residue (Thr669) within the JM domain is phosphorylated by the mitogen-activated protein kinase (MAPK). Previous results have shown that CaM interferes with EGFR-induced MAPK activation. If and how phosphorylation of Thr669 affects CaM-EGFR interaction is however not known.In the present study we have used surface plasmon resonance (BIAcore) to study the influence of Thr669 phosphorylation on real time interactions between the intracellular juxtamembrane (JM) domain of EGFR and CaM. The EGFR-JM was expressed as GST fusion proteins in Escherichia coli and phosphorylation was mimicked by generating Glu substitutions of either Thr654 or Thr669. Purified proteins were coupled to immobilized anti-GST antibodies at the sensor surface and increasing concentration of CaM was applied. When mutating Thr654 to Glu654 no specific CaM binding could be detected. However, neither single substitutions of Thr669 (Gly669 or Glu669) nor double mutants Gly654/Gly669 or Gly654/Glu669 influenced the binding of CaM to the EGFR-JM. This clearly shows that PKC may regulate EGF-mediated CaM signalling through phosphorylation of Thr654 whereas phosphorylation of Thr669 seems to play a CaM independent regulatory role. The role of both residues in the EGFR-calmodulin interaction was also studied in silico. Our modelling work supports a scenario where Thr654 from the JM domain interacts with Glu120 in the calmodulin molecule. Phosphorylation of Thr654 or Glu654 substitution creates a repulsive electrostatic force that would diminish CaM binding to the JM domain. These results are in line with the Biacore experiments showing a weak binding of the CaM to the JM domain with Thr654 mutated to Glu. Furthermore, these results provide a hypothesis to how CaM binding to EGFR might both positively and negatively interfere with EGFR-activity.     

Characterization of recombinant camel chymosin reveals superior properties for the coagulation of bovine and camel milk

Enzymatic milk coagulation for cheese manufacturing involves the cleavage of the scissile bond in kappa-casein by an aspartic acid protease. Bovine chymosin is the preferred enzyme, combining a strong clotting activity with a low general proteolytic activity. In the present study, we report expression and enzymatic properties of recombinant camel chymosin expressed in Aspergillus niger. Camel chymosin was shown to have different characteristics than bovine chymosin. Camel chymosin exhibits a 70% higher clotting activity for bovine milk and has only 20% of the unspecific protease activity for bovine chymosin. This results in a sevenfold higher ratio of clotting to general proteolytic activity. The enzyme is more thermostable than bovine chymosin. Kinetic analysis showed that half-saturation is achieved with less than 50% of the substrate required for bovine chymosin and turnover rates are lower. While raw camel milk cannot be clotted with bovine chymosin, a high clotting activity was found with camel chymosin.