Remodeling of lung interstitium but not resistance vessels in canine pacing-induced heart failure.

We previously showed that pacing-induced heart failure in dogs results in an enhancement of pulmonary vascular reactivity. In the present study we hypothesized that enhanced matrix deposition and structural remodeling of lung resistance microvessels would underlie these functional changes. Using biochemical measures, we found no difference in the normalized lung content of hyaluronan, uronic acid, and collagen between control dogs and dogs paced for 1 mo, although lung dry weight and noncollagen protein content increased significantly in the paced group (P < 0.05). From separate Formalin-fixed lung lobes, 5-microm frozen sections were prepared and stained with Masson's trichrome, and vascular structure was evaluated using standard morphometric techniques. When perivascular fluid cuffs were excluded from the measure of wall thickness, collagen and media volume fractions in any size range did not differ between paced and control groups. Similarly, in the paced group, medial thickness in <400-microm arterial or venular microvessels did not vary significantly from that in the controls. In contrast, the relationship of interstitial fluid pressure to lung water was significantly shifted to the right in the paced group, such that normal tissue pressures were observed, despite the increased water content. We conclude that although 1 mo of pacing-induced heart failure results in altered interstitial function, the attendant pulmonary hypertension and/or hormonal responses are insufficient to induce medial hypertrophy or other remodeling of the extra-alveolar microvasculature.     

Integrated pipeline for mass spectrometry-based discovery and confirmation of biomarkers demonstrated in a mouse model of breast cancer

Despite their potential to impact diagnosis and treatment of cancer, few protein biomarkers are in clinical use. Biomarker discovery is plagued with difficulties ranging from technological (inability to globally interrogate proteomes) to biological (genetic and environmental differences among patients and their tumors). We urgently need paradigms for biomarker discovery. To minimize biological variation and facilitate testing of proteomic approaches, we employed a mouse model of breast cancer. Specifically, we performed LC-MS/MS of tumor and normal mammary tissue from a conditional HER2/Neu-driven mouse model of breast cancer, identifying 6758 peptides representing >700 proteins. We developed a novel statistical approach (SASPECT) for prioritizing proteins differentially represented in LC-MS/MS datasets and identified proteins over- or under-represented in tumors. Using a combination of antibody-based approaches and multiple reaction monitoring-mass spectrometry (MRM-MS), we confirmed the overproduction of multiple proteins at the tissue level, identified fibulin-2 as a plasma biomarker, and extensively characterized osteopontin as a plasma biomarker capable of early disease detection in the mouse. Our results show that a staged pipeline employing shotgun-based comparative proteomics for biomarker discovery and multiple reaction monitoring for confirmation of biomarker candidates is capable of finding novel tissue and plasma biomarkers in a mouse model of breast cancer. Furthermore, the approach can be extended to find biomarkers relevant to human disease.     

Proteinase activity in Ascaris suum eggs, hatching fluid, and excretions-secretions.

Hatching fluid and the excretions and secretions (E.S.) of hatched larvae of Ascaris suum revealed proteinase activity when assayed by 2 different procedures employing collagen or casein as substrates. Both assays apparently detected similar levels of proteinase activity in hatching fluid and E.S. of hatched larvae. The Anson (casein substrate) assay worked best in 0.05 M phosphate buffer, pH 8.0. The Azocoll (collagen substrate) assay worked best in 0.05 M borate buffer at pH 8.8. Azocoll assays done at temperatures ranging from 25 to 65 C revealed maximal proteinase activity at 55 C. Analysis of hatching fluid from 18-, 21-, and 28-day-old embryos and of extracts from sonicated 0- to 28-day-old developmental stages showed that proteinase activity increased markedly 18 days after embryonation had begun. Prior to the 18th day of embryonation proteinase levels were relatively low.     

Hepatic insulin resistance in ob/ob mice involves increases in ceramide,aPKC activity,and selective impairment of Akt-dependent FoxO1 phosphorylation

Pathogenesis of insulin resistance in leptin-deficient ob/ob mice is obscure. In another form of diet-dependent obesity, high-fat-fed mice, hepatic insulin resistance involves ceramide-induced activation of atypical protein kinase C (aPKC), which selectively impairs protein kinase B (Akt)-dependent forkhead box O1 protein (FoxO1) phosphorylation on scaffolding protein, 40 kDa WD(tryp-x-x-asp)-repeat propeller/FYVE protein (WD40/ProF), thereby increasing gluconeogenesis. Resultant hyperinsulinemia activates hepatic Akt and mammalian target of rapamycin C1, and further activates aPKC; consequently, lipogenic enzyme expression increases, and insulin signaling in muscle is secondarily impaired. Here, in obese minimally-diabetic ob/ob mice, hepatic ceramide and aPKC activity and its association with WD40/ProF were increased. Hepatic Akt activity was also increased, but Akt associated with WD40/ProF was diminished and accounted for reduced FoxO1 phosphorylation and increased gluconeogenic enzyme expression. Most importantly, liver-selective inhibition of aPKC decreased aPKC and increased Akt association with WD40/ProF, thereby restoring FoxO1 phosphorylation and reducing gluconeogenic enzyme expression. Additionally, lipogenic enzyme expression diminished, and insulin signaling in muscle, glucose tolerance, obesity, hepatosteatosis, and hyperlipidemia improved. In conclusion, hepatic ceramide accumulates in response to CNS-dependent dietary excess irrespective of fat content; hepatic insulin resistance is prominent in ob/ob mice and involves aPKC-dependent displacement of Akt fromWD40/ProF and subsequent impairment of FoxO1 phosphorylation and increased expression of hepatic gluconeogenic and lipogenic enzymes; and hepatic alterations diminish insulin signaling in muscle.     

Peanut-Cotton Rotations for the Management of Meloidogyne arenaria

The efficacy of ''Deltapine 90'' cotton (Gossypium hirsutum) in rotation with ''Florunner'' peanut (Arachis hypogaea) for the management of Meloidogyne arenaria was studied for 2 years in a field in southeastern Alabama. In 1985, M. arenaria juvenile populations in plots with cotton were 98% lower than in plots with peanut. Peanut and cotton yields were increased by treatment with aldicarb (3.3 kg a.i./ha in a 20-cm-band) in 1985 but not in 1986. In 1986, peanut yields were highest and M. arenaria juvenile populations in soil were lowest in plots that had cotton the previous year. In 1986, numbers of M. arenaria juveniles in plots with peanut both years were reduced by treatment with aldicarb to levels found in plots with cotton-peanut rotation. The use of aldicarb in peanut following cotton similarly treated reduced the incidence of southern blight (Sclerotium rolfsii). Cotton-peanut is a good rotation for the management of M. arenaria and to increase peanut yields without the use of nematicides.     

Postexercise muscle and liver glycogen metabolism in male and female rats

Male and female Wistar rats were run for 5 min at 1.7 mph at a 17% grade to determine whether a sex difference exists in the rate of glycogen resynthesis during recovery in fast-twitch red muscle, fast-twitch white muscle, and liver. Rats were killed at one of three time points: immediately after the exercise bout, and at 1 or 4 h later. Males had significantly higher resting muscle glycogen levels (P less than 0.05). Exercise resulted in significant glycogen depletion in both sexes (P less than 0.01). Males utilized approximately 50% more glycogen during the exercise bout than females (P less than 0.05). During the food-restricted 4-h recovery period, muscle glycogen was repleted significantly during the 1st h (P less than 0.05). Liver glycogen was not depleted as a result of the exercise bout, but fell during the first h of recovery (P less than 0.05) and remained low during the subsequent 3 h. The greater glycogen utilization in red and white fast-twitch muscle during exercise by males could represent a true sex difference but could also be attributable in part to the males having performed more work as a result of 20% greater body mass. We conclude that no sex difference was observed in the rates of muscle glycogen repletion after exercise or in liver glycogen metabolism during and after exercise, and rapid postexercise muscle glycogen repletion occurred at a time of accelerated liver glycogen depletion.