Effect of chronic ethanol administration on the uptake and degradation of asialoglycoproteins by the perfused rat liver

We have shown previously reduced binding, internalization, degradation and receptor-ligand dissociation during receptor-mediated endocytosis (RME) of 125I-asialoorosomucoid (ASOR) by hepatocytes isolated from rats fed ethanol for 4-6 weeks. In the present study, we investigated the effect of ethanol feeding on RME by using the intact perfused liver as a model. Male, Sprague-Dawley rats were fed a liquid diet containing either ethanol (36% of calories) or isocaloric carbohydrate. Receptor-mediated endocytosis of 125I-ASOR was then examined over a time course of perfusion. In all cases, clearance of the labeled glycoprotein was followed by a slower but steady appearance of acid-soluble products in the medium. Ethanol-fed animals had a significantly (P less than 0.01) slower rate of clearance of the labeled ligand from the circulating perfusate than did control animals. Impairment of ASOR surface binding and degradation in ethanol-fed animals was also demonstrated in this model. When we examined the subcellular distribution of labeled ligand after various times of perfusion, we found that in control livers, a shift of radiolabeled ligand from the subcellular fractions containing endosomes and plasma membranes to fractions containing lysosomes occurred, while significantly less ligand was shifted to the lysosomes of ethanol-treated rats. These results show that ethanol administration inhibits RME of ASOR in the isolated perfused liver model, thus confirming our earlier reported defects in isolated hepatocytes. In addition, transport of ligand along the intracellular RME pathway was also shown to be altered by ethanol treatment as indicated by the impaired movement of ASOR from the endosomal to the lysosomal compartment.     

Effects of halothane, sevoflurane and desflurane on the force-frequency relation in the dog heart in vivo

PURPOSE: Frequency potentiation is the increase in force of contraction induced by an increased heart rate (HR). This positive staircase phenomenon has been attributed to changes in Ca2+ entry and loading of intracellular Ca2+ stores. Volatile anesthetics interfere with Ca2+ homeostasis of cardiomyocytes. We hypothesized that frequency potentiation is altered by volatile anesthetics and investigated the influence of halothane (H), sevoflurane (S) and desflurane (D) on the positive staircase phenomenon in dogs in vivo. METHODS: Dogs were chronically instrumented for measurement of left ventricular (LV) pressure and cardiac output. Heart rate was increased by atrial pacing from 120 to 220 beats x min(-1) and the LV maximal rate of pressure increase (dP/dt(max)) was determined as an index of myocardial performance. Measurements were performed in conscious dogs and during anesthesia with 1.0 minimal alveolar concentrations of each of the three inhaled anesthetics. RESULTS: Increasing HR from 120 to 220 beats x min(-1) increased dP/dt(max) from 3394 +/- 786 (mean +/- SD) to 3798 +/- 810 mmHg sec(-1) in conscious dogs. All anesthetics reduced dP/dt(max) during baseline (at 120 beats x min(-1): H, 1745 +/- 340 mmHg x sec(-1); S, 1882 +/- 418; D, 1928 +/- 454, all P < 0.05 vs awake) but did not influence the frequency potentiation of dP/dt(max) (at 220 beats x min(-1): H, 1981 +/- 587 mmHg x sec(-1); S, 2187 +/- 787; D, 2307 +/- 691). The slope of the regression line correlating dP/dt(max) and HR was not different between awake and anesthetized dogs. Increasing HR did not influence cardiac output in awake or anesthetized dogs. CONCLUSION: These results indicate that volatile anesthetics do not alter the force-frequency relation in dogs in vivo.     

Rate of nitrous oxide exchange across the middle ear mucosa in monkeys before and after blockage of the mastoid antrum.

OBJECTIVES: We tested the hypothesis that mastoid volume buffers the rate of change in middle ear pressure caused by transmucosal, inert gas exchange. STUDY DESIGN: Twelve monkeys were randomly assigned to group 1 or group 2. Right ears of group 1 had sham surgery and of group 2 had obstruction of the mastoid antrum. Before and after surgery, the time constant for transmucosal N(2)O exchange was estimated from N(2)O breathing experiments. The hypothesis predicts that the postoperative time constant measured for right ears of group 2 but not group 1 is greater than that measured before surgery. RESULTS: Mastoid antrum block significantly decreased right middle ear volume but did not affect the time constant for transmucosal N(2)O exchange. CONCLUSION: A mastoid gas-reserve function is not supported by the experimental data. SIGNIFICANCE: These results for monkeys and the theory developed to explain the effect of mastoid volume on transmucosal inert gas exchange suggest that the results for previous experiments in humans interpreted as evidencing a mastoid gas-reserve function are consistent with alternative explanations.     

Testosterone suppresses protective responses of the liver to blood-stage malaria

Testosterone induces a lethal outcome in otherwise self-healing blood-stage malaria caused by Plasmodium chabaudi. Here, we examine possible testosterone effects on the antimalaria effectors spleen and liver in female C57BL/6 mice. Self-healing malaria activates gating mechanisms in the spleen and liver that lead to a dramatic reduction in trapping activity, as measured by quantifying the uptake of 3-mum-diameter fluorescent polystyrol particles. However, testosterone delays malaria-induced closing of the liver, but not the spleen. Coincidently, testosterone causes an approximately 3- to 28-fold depression of the mRNA levels of nine malaria-responsive genes, out of 299 genes tested, only in the liver and not in the spleen, as shown by cDNA arrays and Northern blotting. Among these are the genes encoding plasminogen activator inhibitor (PAI1) and hydroxysteroid sulfotransferase (STA2). STA2, which detoxifies bile acids, is suppressed 10-fold by malaria and an additional 28-fold by testosterone, suggesting a severe perturbation of bile acid metabolism. PAI1 is protective against malaria, since disruption of the PAI1 gene results in partial loss of the ability to control the course of P. chabaudi infections. Collectively, our data indicate that the liver rather than the spleen is a major target organ for testosterone-mediated suppression of resistance against blood-stage malaria.     

A unique exonic splice enhancer mutation in a family with X-linked mental retardation and epilepsy points to a novel role of the renin receptor

The renin-angiotensin system (RAS) is essential for blood pressure control and water-electrolyte balance. Until the discovery of the renin receptor, renin was believed to be mainly a circulating enzyme with a unique function, the cleavage of angiotensinogen. We report a unique mutation in the renin receptor gene (ATP6AP2) present in patients with X-linked mental retardation and epilepsy (OMIM no. 300423), but absent in 1200 control X-chromosomes. A silent mutation (c.321C>T, p.D107D) residing in a putative exonic splicing enhancer site resulted in inefficient inclusion of exon 4 in 50% of renin receptor mRNA, as demonstrated by quantitative RT-PCR. Analysis of membrane associated-receptor molecular forms showed the presence of full-length and truncated proteins in the patient. Functional analysis demonstrated that the mutated receptor could bind renin and increase renin catalytic activity, similar to the wild-type receptor, but resulted in a modest and reproducible impairment of ERK1/2 activation. Thus, our findings confirm the importance of the RAS in cognitive processes and indicate a novel specific role for the renin receptor in cognitive functions and brain development.     

Early detection and successful therapy of fulminant chlamydia pneumoniae myocarditis

We report a case involving a young female patient with Chlamydia pneumoniae myocarditis who required assist device therapy for acute heart failure. Early diagnosis was provided by endomyocardial biopsy, and tailored antibiotic therapy facilitated quick recovery of myocardial function. This is the first case to report detecting C pneumoniae as the pathogen responsible for fulminant myocarditis while the patient was still alive and to report long-term follow-up data.     

T cell epitope spreading to myelin oligodendrocyte glycoprotein in HLA-DR4 transgenic mice during experimental autoimmune encephalomyelitis

Epitope spreading has been implicated in the pathogenesis of experimental autoimmune encephalomyelitis (EAE) and human multiple sclerosis (MS). T cell epitope spreading has been demonstrated in rodents for myelin basic protein (MBP) and proteolipid protein (PLP) determinants, but not for myelin oligodendrocyte glycoprotein (MOG), another important myelin antigen. Moreover, the role of human autoimmunity-associated MHC molecules in epitope spreading, including HLA-DR2 and DR4, has not been formally examined. To address these questions, we investigated epitope spreading to MOG determinants in HLA-DR4 (DRB1*0401) transgenic mice during EAE. The data show that upon induction of EAE in HLA-DR4 transgenic mice with the immunodominant HLA-DR4-restricted MOG peptide 97-108 (MOG(97-108); TCFFRDHSYQEE), the T cell response diversifies over time to MOG(181-200) (core: MOG(183-191); FVIVPVLGP) and MBP. The spreading epitope MOG(181-200) binds with high affinity to HLA-DRB1*0401 and is presented by human HLA-DRB1*0401+antigen presenting cells. Moreover, this epitope is encephalitogenic in HLA-DRB1*0401 transgenic mice. This study demonstrates intra- and intermolecular epitope spreading to MOG and MBP in "humanized" HLA-DR4 transgenic mice.     

Resolution and conservation of mismatches in DNA end joining

DNA end joining is a major pathway for the elimination of double-strandbreaks from chromosomal DNA of higher eucaryotic cells. Extractsof Xenopus laevis eggs rejoin such breaks even when their shortsingle-stranded termini are expected to form imperfectly matchedoverlaps. However, end-joined products cloned in Escherichiacoli, necessarily give rise to perfectly matched products. Thereforeit has not been possible to determine whether the end joiningprocess creates mismatched products, perfectly matched (resolved)products or both. To investigate whether mismatch resolutionwas the result of the X. laevis end joining process or of activitiesof the bacterial host we used denaturing gradient gel electrophoresisto analyse joined products. We found that the end joining processdoes include mismatch resolution, the degree of which varieswith regard to the nature of the original overlap structure.Mismatches 3' to a gap are completely resolved, mismatches 3'to a nick and 5' to a nick or gap are resolved to some extentbut are generally conserved. Mismatches between base matchesare always conserved. These findings suggest competing processesof ligation, DNA fill-in synthesis or exonucleolytic excisionof mismatched bases next to a gap or nick. At mismatches 3'to a nick the probability of ligation is greater than that ofexcision while at mismatches 3' to a gap the probability ofexcision is greater than elongation of a given mismatch. Atmismatches 5' to nicks or gaps it appears that ligation or elongationand ligation, respectively, are the most probable pathways butproducts resulting from mismatch excision, elongation and ligationare also detected. ⁵To whom correspondence should be addressed