Studien über die von einzelnen Organen hervorgebrachten Substanzen mit spezifischer Wirkung

Ohne ZusammenfassungEmil Abderhalden, Dieses Arch. 162, 99. 1915;176, 236.1919. — Emil Abderhalden und Ernest Gellhorin, Ebenda183, 1920.     

Enzymatic and functional correction along with long-term enzyme secretion from transduced bone marrow hematopoietic stem/progenitor and stromal cells derived from patients with Fabry disease.

Fabry disease is a lysosomal storage disorder that is due to a deficiency in alpha-galactosidase A (alpha-gal A). Previously we have shown that a recombinant retrovirus synthesized for the transfer of the human alpha-gal A coding sequence was able to engineer enzymatic correction of the hydrolase deficiency in fibroblasts and lymphoblasts from Fabry patients. The corrected cells secreted alpha-gal A that was taken up and utilized by uncorrected bystander cells, thus demonstrating metabolic cooperativity. In separate experiments we used transduced murine bone marrow cells and successfully tested and quantitated this phenomenon in vivo. In the present studies, which were designed to bring this therapeutic approach closer to clinical utility, we establish that cells originating from the bone marrow of numerous Fabry patients and normal volunteers can be effectively transduced and that these target cells demonstrate metabolic cooperativity. Both isolated CD34+-enriched cells and long-term bone marrow culture cells, including nonadherent hematopoietic cells and adherent stromal cells, were transduced. The transferred gene generates increased intracellular alpha-gal A enzyme activity in these cells. Further, it causes functional correction of lipid accumulation and provides for long-term alpha-gal A secretion. Collectively, these results indicate that a multifaceted gene transfer approach to bone marrow cells may be of therapeutic benefit for patients with Fabry disease.     

An approach to teaching the introduction to clinical medicine.

An approach to teaching the introduction of clinical medicine to medical students is described. It is based on (A) gradual exposure to patients and step-by-step introduction to the objectives and techniques of history taking, with emphasis on training for symptom identification; (B) early exposure to the problem-oriented approach; (C) early diagnostic hypothesis generation; and (D) self-directed learning. The objectives of the proposed approach are to teach students to communicate with patients, to teach the skills of history taking and physical examination, and to introduce the problem-solving strategies that characterize the diagnostic reasoning of experienced physicians.     

Impaired motor coordination and Purkinje cell excitability in mice lacking calretinin

In the cerebellum, the parallel fiber-Purkinje cell synapse can undergo long-term synaptic plasticity suggested to underlie motor learning and resulting from variations in intracellular calcium concentration ([Ca2+]i). Ca2+ binding proteins are enriched in the cerebellum, but their role in information processing is not clear. Here, we show that mice deficient in calretinin (Cr-/-) are impaired in tests of motor coordination. An impairment in Ca2+ homeostasis in Cr-/- Purkinje cells was supported by the high Ca2+-saturation of calbindin-D28k in these cells. The firing behavior of Purkinje cells is severely affected in Cr-/- alert mice, with alterations of simple spike firing rate, complex spike duration, and simple spike pause. In contrast, in slices, transmission at parallel fiber- or climbing fiber-Purkinje cell synapses is unaltered, indicating that marked modifications of the firing behavior in vivo can be undetectable in slice. Thus, these results show that calretinin plays a major role at the network level in cerebellar physiology.     

Allosteric interactions between agonists and antagonists within the adenosine A2A receptor-dopamine D2 receptor heterotetramer

Adenosine A_2A receptor (A_2AR)-dopamine D₂ receptor (D₂R) heteromers are key modulators of striatal neuronal function. It has been suggested that the psychostimulant effects of caffeine depend on its ability to block an allosteric modulation within the A_2AR-D₂R heteromer, by which adenosine decreases the affinity and intrinsic efficacy of dopamine at the D₂R. We describe novel unsuspected allosteric mechanisms within the heteromer by which not only A_2AR agonists, but also A_2AR antagonists, decrease the affinity and intrinsic efficacy of D₂R agonists and the affinity of D₂R antagonists. Strikingly, these allosteric modulations disappear on agonist and antagonist coadministration. This can be explained by a model that considers A_2AR-D₂R heteromers as heterotetramers, constituted by A_2AR and D₂R homodimers, as demonstrated by experiments with bioluminescence resonance energy transfer and bimolecular fluorescence and bioluminescence complementation. As predicted by the model, high concentrations of A_2AR antagonists behaved as A_2AR agonists and decreased D₂R function in the brain.Most evidence indicates that G protein-coupled receptors (GPCRs) form homodimers and heteromers. Homodimers seem to be a predominant species, and oligomeric entities can be viewed as multiples of dimers (1). It has been proposed that GPCR heteromers are constituted mainly by heteromers of homodimers (1, 2). Allosteric mechanisms determine a multiplicity of unique pharmacologic properties of GPCR homodimers and heteromers (1, 3). First, binding of a ligand to one of the receptors in the heteromer can modify the affinity of ligands for the other receptor (1, 3, 4). The most widely reproduced allosteric modulation of ligand-binding properties in a GPCR heteromer is the ability of adenosine A_2A receptor (A_2AR) agonists to decrease the affinity of dopamine D₂ receptor (D₂R) agonists in the A_2AR-D₂R heteromer (5). A_2AR-D₂R heteromers have been revealed both in transfected cells (6, 7), striatal neurons in culture (6, 8) and in situ, in mammalian striatum (9, 10), where they play an important role in the modulation of GABAergic striatopallidal neuronal function (9, 11).In addition to ligand-binding properties, unique properties for each GPCR oligomer emerge in relation to the varying intrinsic efficacy of ligands for different signaling pathways (1–3). Intrinsic efficacy refers to the power of the agonist to induce a functional response, independent of its affinity for the receptor. Thus, allosteric modulation of an agonist can potentially involve changes in affinity and/or intrinsic efficacy (1, 3). This principle can be observed in the A_2AR-D₂R heteromer, where a decrease in D₂R agonist affinity cannot alone explain the ability of an A_2AR agonist to abolish the decreased excitability of GABAergic striatopallidal neurons induced by high concentration of a D₂R agonist (9), which should overcome the decrease in affinity. Furthermore, a differential effect of allosteric modulations of different agonist-mediated signaling responses (i.e., functional selectivity) can occur within GPCR heteromers (1, 2, 8). Again, the A_2AR-D₂R heteromer provides a valuable example. A recent study has shown that different levels of intracellular Ca²⁺ exert different modulations of A_2AR-D₂R heteromer signaling (8). This depends on the ability of low and high Ca²⁺ to promote a selective interaction of the heteromer with different Ca²⁺-binding proteins, which differentially modulate allosteric interactions in the heteromer (8).It has been hypothesized that the allosteric interactions between A_2AR and D₂R agonists within the A_2AR-D₂R heteromer provide a mechanism responsible not only for the depressant effects of A_2AR agonists, but also for the psychostimulant effects of adenosine A_2AR antagonists and the nonselective adenosine receptor antagonist caffeine (9, 11, 12), with implications for several neuropsychiatric disorders (13). In fact, the same mechanism has provided the rationale for the use of A_2AR antagonists in patients with Parkinson’s disease (13, 14). The initial aim of the present study was to study in detail the ability of caffeine to counteract allosteric modulations between A_2AR and D₂R agonists (affinity and intrinsic efficacy) within the A_2AR-D₂R heteromer. Unexpectedly, when performing control radioligand-binding experiments, not only an A_2AR agonist, but also caffeine, significantly decreased D₂R agonist binding. However, when coadministered, the A_2AR agonist and caffeine co-counteracted their ability to modulate D₂R agonist binding. By exploring the molecular mechanisms behind these apparent inconsistencies, the present study provides new insight into the quaternary structure and function of A_2AR-D₂R heteromers.     

N-formylmethionyl peptides as chemoattractants for leucocytes.

Leucocytes such as neutrophils are attracted by N-formylmethionine, but not by methionine. Di- and tripeptides containing formylmethionine are strong attractants for both neutrophils and macrophages, whereas the corresponding nonacylated compounds are not chemotactic. The formylated peptides may be related to an incompletely characterized chemotactic material normally produced by bacteria which attract the same animal cells.     

A double-blind,placebo-controlled trial of triheptanoin in adult polyglucosan body disease and open-label,long-term outcome

Background

Adult polyglucosan body disease (APBD) is a progressive neurometabolic disorder caused by a deficiency of glycogen branching enzyme. We tested the efficacy of triheptanoin as a therapy for patients with APBD based on the hypothesis that decreased glycogen degradation leads to brain energy deficit.

Methods and results

This was a two-site, randomized crossover trial of 23 patients (age 35–73 years; 63% men) who received triheptanoin or vegetable oil as placebo. The trial took place over 1 year and was followed by a 4-year open-label phase. Generalized linear mixed models were used to analyze this study. At baseline, using the 6-min walk test, patients could walk a mean of 389 ± 164 m (range 95–672; n = 19), highlighting the great clinical heterogeneity of our cohort. The overall mean difference between patients on triheptanoin versus placebo was 6 m; 95% confidence interval (CI) ?11 to 22; p = 0.50. Motion capture gait analysis, gait quality, and stair climbing showed no consistent direction of change. All secondary endpoints were statistically nonsignificant after false discovery rate adjustment. Triheptanoin was safe and generally well tolerated. During the open-label phase of the study, the most affected patients at baseline kept deteriorating while mildly disabled patients remained notably stable up to 4 years.

Conclusions

We cannot conclude that triheptanoin was effective in the treatment of APBD over a 6-month period, but we found it had a good safety profile. This study also emphasizes the difficulty of conducting trials in very rare diseases presenting with a wide clinical heterogeneity. ClinicalTrials.gov Identifier: NCT00947960.