Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts

The concept of regenerating diseased myocardium by implantation of tissue-engineered heart muscle is intriguing, but convincing evidence is lacking that heart tissues can be generated at a size and with contractile properties that would lend considerable support to failing hearts. Here we created large (thickness/diameter, 1-4 mm/15 mm), force-generating engineered heart tissue from neonatal rat heart cells. Engineered heart tissue formed thick cardiac muscle layers when implanted on myocardial infarcts in immune-suppressed rats. When evaluated 28 d later, engineered heart tissue showed undelayed electrical coupling to the native myocardium without evidence of arrhythmia induction. Moreover, engineered heart tissue prevented further dilation, induced systolic wall thickening of infarcted myocardial segments and improved fractional area shortening of infarcted hearts compared to controls (sham operation and noncontractile constructs). Thus, our study provides evidence that large contractile cardiac tissue grafts can be constructed in vitro, can survive after implantation and can support contractile function of infarcted hearts.     

Impaired cerebral cortex development and blood pressure regulation in FGF-2-deficient mice.

Fibroblast growth factor-2 (FGF-2) has been implicated in various signaling processes which control embryonic growth and differentiation, adult physiology and pathology. To analyze the in vivo functions of this signaling molecule, the FGF-2 gene was inactivated by homologous recombination in mouse embryonic stem cells. FGF-2-deficient mice are viable, but display cerebral cortex defects at birth. Bromodeoxyuridine pulse labeling of embryos showed that proliferation of neuronal progenitors is normal, whereas a fraction of them fail to colonize their target layers in the cerebral cortex. A corresponding reduction in parvalbumin-positive neurons is observed in adult cortical layers. Neuronal defects are not limited to the cerebral cortex, as ectopic parvalbumin-positive neurons are present in the hippocampal commissure and neuronal deficiencies are observed in the cervical spinal cord. Physiological studies showed that FGF-2-deficient adult mice are hypotensive. They respond normally to angiotensin II-induced hypertension, whereas neural regulation of blood pressure by the baroreceptor reflex is impaired. The present genetic study establishes that FGF-2 participates in controlling fates, migration and differentiation of neuronal cells, whereas it is not essential for their proliferation. The observed autonomic dysfunction in FGF-2-deficient adult mice uncovers more general roles in neural development and function.     

A new technique for determining the hydride transfer stereospecificity of nicotinamide adenine dinucleotide-linked oxidoreductases: Electron impact and field desorption mass spectrometry

The stereospecificity of nicotinamide adenine dinucleotide-linked oxidoreductases can be determined directly without additional purification and with high sensitivity by low-resolution electron impact mass spectrometry and low-resolution field desorption mass spectrometry. The ease and reliability of this new technique is demonstrated by studying the hydride transfer stereospecificity of lactate dehydrogenase from pig heart (known to be A-specific) and glutamate dehydrogenase from beef liver (known to be B-specific). In addition the stereospecificity of glutamate dehydrogenases from duckweed (Lemna minor) was found to be B-specific. The sample quantity of the unpurified products needed is about 1 μg. The time required for one mass spectrometric determination was about 30 min.     

Increased synaptic microtubules and altered synapse development inDrosophila sec8 mutants

Background  

Sec8 is highly expressed in mammalian nervous systems and has been proposed to play a role in several aspects of neural development and function, including neurite outgrowth, calcium-dependent neurotransmitter secretion, trafficking of ionotropic glutamate receptors and regulation of neuronal microtubule assembly. However, these models have never been testedin vivo. Nervous system development and function have not been described after mutation ofsec8 in any organism.     

Role of mitochondrial glutamate dehydrogenase in the reassimilation of ammonia produced by glycine serine transformation

The ability of isolated pea-shoot mitochondria conditioned to incorporate ammonia into glutamate to reassimilate endogenously produced ammonia from glycine transformation was investigated. In the presence of 1 mM to 20 mM glycine less than 15% of the ammonia liberated was found to be incorporated into glutamate. Thus, a prominent role of mitochondrial glutamate dehydrogenase in the reassimilation of intramitochondrially produced ammonia can be excluded.Abbreviation GDH Glutamate dehydrogenase (L-glutamate: NAD⁺ oxidoreductase (deaminating), EC 1.4.1.2)     

Revealing biases inherent in recombination protocols

Background  

The recombination of homologous genes is an effective protein engineering tool to evolve proteins. DNA shuffling by gene fragmentation and reassembly has dominated the literature since its first publication, but this fragmentation-based method is labor intensive. Recently, a fragmentation-free PCR based protocol has been published, termed recombination-dependent PCR, which is easy to perform. However, a detailed comparison of both methods is still missing.