Immunohistochemical Demonstration of Leptin in Pancreatic Islets of Non-Obese Diabetic and CD-1 mice: Co-localization in Glucagon Cells and its Attenuation at the Onset of Diabetes

Leptin is a 16 kD polypeptide hormone produced predominantly by white adipose tissue and exerts profound effects on food intake and energy balance. More recent studies have shown extra sites of leptin production in human and rodent tissues and have ascribed additional roles for the hormone, e.g., in immune and reproductive functions. A role for the hormone has also been implicated in insulin-dependent diabetes mellitus in the non-obese diabetic (NOD) mouse. However, whether leptin originates from islet cells of the mouse is not known. Here dual-label immunohistochemistry was employed to examine leptin expression in islet cells, and its distribution and cellular sources in pancreatic sections of female NOD/Ak and CD-1 mice of various ages. For comparison, leptin immunolabelling was examined in adult pancreatic sections from male NOD/Ak CD-1, Balb/c and FVB/N mice and female severe combined immunodeficient CB. 17 mice. Pancreatic tissues from adult female guinea pig, sheep and cattle and neonatal pigs were also studied. Our results show that in the day 1 NOD and CD-1 mice, leptin immunolabelling was observed in selective glucagon cells within the developing islets while at days 15 and 22, it became more intense and co-incident. This pattern of staining was maintained at days 40, 90, 150 and 250. In the female NOD mouse, leptin was absent in intra-islet immune cells. Its expression was variable in islets from male NOD and CD-1 mice. In spontaneously diabetic female NOD mice and following acceleration of diabetes with cyclophosphamide, despite the persistence of strong immunolabelling for glucagon in the re-distributed alpha cells, leptin expression was either absent, diminished or present in only a proportion of alpha cells. The reduction in leptin labelling was often associated with diabetic islets which had insulitis in association with only a small number of residual beta cells. Leptin expression was absent in guinea pig, ovine, bovine and neonatal porcine islet cells, despite the expression of intensely labelled glucagon cells. The present results demonstrate leptin co-localization in glucagon cells of the mouse islet. Its expression diminishes in the presence of inadequate insulin. Leptin produced within the mouse islet may have bi-directional influences on leptin and insulin regulation and may play local functions in islet development and metabolism.     

Relative Ability of Orally Administered Lactobacillus murinus To Predominate and Persist in the Porcine Gastrointestinal Tract

Five porcine-derived Lactobacillus or Pediococcus isolates administered to pigs (n = 4), either singly or as a combination at ~10¹⁰ CFU per day varied with respect to intestinal survival and persistence. Two Lactobacillus murinus strains survived best and were excreted at ~10⁷ to 10⁸ CFU/g of feces. In contrast, Pediococcus pentosaceus DPC6006 had the lowest fecal count at ~10⁵ CFU/g and was excreted at a significantly lower level than both L. murinus strains. Fecal L. murinus DPC6003 counts were also significantly higher than both Lactobacillus salivarius DPC6005 and Lactobacillus pentosus DPC6004 (~10⁶ CFU/g). The L. murinus strains persisted for at least 9 days postadministration in both the feces and the cecum. Animals fed a combination of all five strains excreted ~10⁷ CFU of the administered strains/g, with L. murinus predominating, as determined by randomly amplified polymorphic DNA PCR. Postadministration, variation was observed between animals fed the strain combination, but in general, L. murinus DPC6002 and DPC6003 and L. pentosus DPC6004 predominated in the feces and the cecum while P. pentosaceus DPC6006 was detected only in the cecum. Fifteen days after the start of culture administration, mean fecal Enterobacteriaceae counts were significantly lower in some of the treatment groups. In addition, when mean preadministration counts were compared with those obtained after 21 days of culture administration, Enterobacteriaceae counts were reduced by ~87 to 98% in pigs fed L. salivarius DPC6005, P. pentosaceus DPC6006, L. pentosus DPC6004, and the culture mix. In conclusion, the porcine intestinal isolates have potential as probiotic feed additives for pigs, with differences in strain performance highlighting the advantages of using culture combinations.     

Development of an assay to screen for inhibitors of tau phosphorylation by cdk5

A high-throughput assay for tau phosphorylation by cdk5/p25 is described. Full-length recombinant tau was used as a substrate in the presence of saturating adenosine triphosphate (ATP). Using PHF-1, an antibody directed specifically against 2 tau phosphorylation epitopes (serine 396 and serine 404), an enzyme-linked immunosorbent assay (ELISA)-based colorimetric assay was formatted in 384-well plates. The assay was validated by measuring kinetic parameters for cdk5/p25 catalysis and known inhibitors. Rate constants for the site-specific phosphorylations at the PHF-1 epitopes were determined and suggested preferential phosphorylation at these sites. The performance of this assay in a high-throughput format was demonstrated and used to identify inhibitors of tau phosphorylation at specific epitopes phosphorylated by cdk5/p25.     

Modelling the Species Distribution of Flat-Headed Cats (Prionailurus planiceps), an Endangered South-East Asian Small Felid

Background

The flat-headed cat (Prionailurus planiceps) is one of the world''s least known, highly threatened felids with a distribution restricted to tropical lowland rainforests in Peninsular Thailand/Malaysia, Borneo and Sumatra. Throughout its geographic range large-scale anthropogenic transformation processes, including the pollution of fresh-water river systems and landscape fragmentation, raise concerns regarding its conservation status. Despite an increasing number of camera-trapping field surveys for carnivores in South-East Asia during the past two decades, few of these studies recorded the flat-headed cat.

Methodology/Principal Findings

In this study, we designed a predictive species distribution model using the Maximum Entropy (MaxEnt) algorithm to reassess the potential current distribution and conservation status of the flat-headed cat. Eighty-eight independent species occurrence records were gathered from field surveys, literature records, and museum collections. These current and historical records were analysed in relation to bioclimatic variables (WorldClim), altitude (SRTM) and minimum distance to larger water resources (Digital Chart of the World). Distance to water was identified as the key predictor for the occurrence of flat-headed cats (>50% explanation). In addition, we used different land cover maps (GLC2000, GlobCover and SarVision LLC for Borneo), information on protected areas and regional human population density data to extract suitable habitats from the potential distribution predicted by the MaxEnt model. Between 54% and 68% of suitable habitat has already been converted to unsuitable land cover types (e.g. croplands, plantations), and only between 10% and 20% of suitable land cover is categorised as fully protected according to the IUCN criteria. The remaining habitats are highly fragmented and only a few larger forest patches remain.

Conclusion/Significance

Based on our findings, we recommend that future conservation efforts for the flat-headed cat should focus on the identified remaining key localities and be implemented through a continuous dialogue between local stakeholders, conservationists and scientists to ensure its long-term survival. The flat-headed cat can serve as a flagship species for the protection of several other endangered species associated with the threatened tropical lowland forests and surface fresh-water sources in this region.     

Electromyography of the anterior temporalis and masseter muscles of owl monkeys (Aotus trivirgatus) and the function of the postorbital septum

Anthropoids and tarsiers are distinguished from all other vertebrates by the possession of a postorbital septum, which is formed by the frontal, alisphenoid, and zygomatic bones. Cartmill [(1980) In: Evolutionary Biology of the New World Monkeys and Continental Drift. New York: Plenum, p 243-274] suggested that the postorbital septum evolved in the stem lineage of tarsiers and anthropoids to insulate the eye from movements arising in the temporal fossa. Ross [(1996) Am J Phys Anthropol 91:305-324] suggested that the septum insulates the orbital contents from incursions by the line of action of the anterior temporal muscles caused by the unique combination of high degrees of orbital frontation and convergence. Both of these hypotheses must explain why insulation of the orbital contents could not be achieved by decreasing the size of the anterior temporal musculature with a corresponding increase in size of the remaining jaw adductors, rather than evolving a postorbital septum. One possibility is that the anterior temporalis is an important contributor to vertically directed bite forces during all biting and chewing activities. Another possibility is that reduction in anterior temporal musculature would compromise the ability to produce powerful bite forces, either at the incisors or along the postcanine toothrow. To evaluate these hypotheses, electromyographic (EMG) recordings were made from the masseter muscle and the anterior and posterior portions of the temporalis muscles of two owl monkeys, Aotus trivirgatus. The EMG data indicate that anterior temporalis activity relative to that of the superficial masseter is lower during incision than mastication. In addition, activity of the anterior temporalis is not consistently higher than the posterior temporalis during incision. The data indicate relatively greater activity of anterior temporalis compared to other muscles during isometric biting on the postcanine toothrow. This may be due to decreased activity in superficial masseter and posterior temporalis, rather than elevated anterior temporalis activity. The anterior temporalis is not consistently less variable in activity than the superficial masseter and posterior temporalis. The EMG data gathered here indicate no reason for suggesting that the anterior temporal muscles in anthropoids are utilized especially for incisal preparation of hard fruits. Maintenance of relatively high EMG activity in anterior temporalis across a wide range of biting behaviors is to be expected in a vertically oriented and rostrally positioned muscle such as this because, compared to the posterior temporalis, superficial masseter and medial pterygoid, it can contribute relatively larger vertical components of force to bites along the postcanine toothrow. The in vivo data do not support this hypothesis, possibly because of effects of bite point and bite force orientation.     

Hyperthermic modification of cyclophosphamide metabolism in rat hepatic microsomes and liver slices

The ability of rat liver microsomes and liver slices to metabolize the antineoplastic compound cyclophosphamide was studied at 37° and at elevated temperatures comparable to those used for human systemic hyperthermic antineoplastic therapy. Temperatures above 40.5° and 41.8° inhibited cyclophospamide metabolism by microsomes and liver slices respectively. Therefore, cyclophosphamide may not be a suitable drug for combination with systemic hyperthermia in cancer therapy.     

Resource tracking in marine parasites: going with the flow?

Understanding how diversity interacts with energy supply is of broad ecological interest. Most studies to date have investigated patterns within trophic levels, reflecting a lack of food webs which include information on energy flow. We added parasites to a published marine energy‐flow food web, to explore whether parasite diversity is correlated with energy flow to host taxa. Parasite diversity was high with 36 parasite taxa affecting 40 of the 51 animal taxa. Adding parasites increased the number of trophic links per species, trophic link strength, connectance, and food chain lengths. There was evidence of an asymptotic relationship between energy flowing through a food chain and parasite diversity, although there were clear outliers. High parasite diversity was associated with host taxa which were highly connected within the food web. This suggests that energy flow through a taxon may favour parasite diversity, up to a maximal value. The evolutionary and energetic basis for that limitation is of key interest in understanding the basis for parasite diversity in natural food webs and thus their role in food web dynamics.     

A landscape approach for identifying potential reestablishment sites for extirpated stream fishes: an example with Arctic grayling (Thymallus arcticus) in Michigan

Hydrobiologia - Habitat degradation combined with climate change increases the threat of extinction for stream fishes. In response to these threats, efforts to reestablish species within formerly...     

A Regional Initiative to Reduce Skin Infections amongst Aboriginal Children Living in Remote Communities of the Northern Territory,Australia

Background

Linked to extreme rates of chronic heart and kidney disease, pyoderma is endemic amongst Aboriginal children in Australia''s Northern Territory (NT). Many of those with pyoderma will also have scabies. We report the results of a community-based collaboration within the East Arnhem Region, which aimed to reduce the prevalence of both skin infections in Aboriginal children.

Methodology/Principal Findings

Commencing September 2004, we conducted an ecological study that included active surveillance for skin infections amongst children aged <15 years in five remote East Arnhem communities over a three year period. Screening was undertaken by trained local community workers, usually accompanied by another project team member, using a standard data collection form. Skin infections were diagnosed clinically with the aid of a pictorial flip chart developed for the purpose. Topical 5% permethrin was provided for age-eligible children and all household contacts whenever scabies was diagnosed, whilst those with pyoderma were referred to the clinic for treatment in accordance with current guidelines. In addition, annual mass scabies treatment (5% permethrin cream) was offered to all community residents in accordance with current guidelines but was not directly observed. Pyoderma and scabies prevalence per month was determined from 6038 skin assessments conducted on 2329 children. Pyoderma prevalence dropped from 46.7% at baseline to a median of 32.4% (IQR 28.9%–41.0%) during the follow-up period – an absolute reduction of 14.7% (IQR 4.7%–16.8%). Compared to the first 18 months of observation, there was an absolute reduction in pyoderma prevalence of 18 cases per 100 children (95%CI −21.0, −16.1, p≤0.001) over the last 18 months. Treatment uptake increased over the same period (absolute difference 13.4%, 95%CI 3.3, 23.6). While scabies prevalence was unchanged, the prevalence of infected scabies (that is with superimposed pyoderma) decreased from 3.7% (95%CI 2.4, 4.9) to 1.5% (95%CI 0.7, 2.2), a relative reduction of 59%.

Conclusion

Although pyoderma prevalence remained unacceptably high, there was a substantial reduction overall with improvements in treatment uptake a critical factor. More acceptable alternatives, such as cotrimoxazole for pyoderma and ivermectin as a community-wide scabicide, warrant further investigation in these settings. We are encouraged by progress made through this work, where local action was led by local community members and primary health care providers with external training and support.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00884728","term_id":"NCT00884728"}}NCT00884728     

beta-Adrenergic receptor-stimulated apoptosis in adult cardiac myocytes involves MMP-2-mediated disruption of beta1 integrin signaling and mitochondrial pathway

Stimulation of -adrenergic receptors (-AR) induces apoptosis in adult rat ventricular myocytes (ARVMs) via the JNK-dependent activation of mitochondrial death pathway. Recently, we have shown that inhibition of matrix metalloproteinase-2 (MMP-2) inhibits -AR-stimulated apoptosis and that the apoptotic effects of MMP-2 are possibly mediated via its interaction with ₁ integrins. Herein we tested the hypothesis that MMP-2 impairs ₁ integrin-mediated survival signals, such as activation of focal adhesion kinase (FAK), and activates the JNK-dependent mitochondrial death pathway. Inhibition of MMP-2 using SB3CT, a selective gelatinase inhibitor, significantly increased FAK phosphorylation (Tyr-397 and Tyr-576). TIMP-2, tissue inhibitor of MMP-2, produced a similar increase in FAK phosphorylation, whereas treatment of ARVMs with purified active MMP-2 significantly inhibited FAK phosphorylation. Inhibition of MMP-2 using SB3CT inhibited -AR-stimulated activation of JNKs and levels of cytosolic cytochrome c. Treatment of ARVMs with purified MMP-2 increased cytosolic cytochrome c release. Furthermore, inhibition of MMP-2 using SB3CT and TIMP-2 attenuated -AR-stimulated decreases in mitochondrial membrane potential. Overexpression of ₁ integrins using adenoviruses expressing the human _1A-integrin decreased -AR-stimulated cytochrome c release and apoptosis. Overexpression of ₁ integrins also inhibited apoptosis induced by purified active MMP-2. These data suggest that MMP-2 interferes with the ₁ integrin survival signals and activates JNK-dependent mitochondrial death pathway leading to apoptosis. matrix metalloproteinases; focal adhesion kinase; c-Jun NH₂-terminal kinase; cytochrome c