Tooth enamel forms in an ephemeral protein matrix where changes in protein abundance, composition and posttranslational modifications are critical to achieve healthy enamel properties. Amelogenin (AMELX) with its splice variants is the most abundant enamel matrix protein, with only one known phosphorylation site at serine 16 shown in vitro to be critical for regulating mineralization. The phosphorylated form of AMELX stabilizes amorphous calcium phosphate, while crystalline hydroxyapatite forms in the presence of the unphosphorylated protein. While AMELX regulates mineral transitions over space and time, it is unknown whether and when un-phosphorylated amelogenin occurs during enamel mineralization. This study aims to reveal the spatiotemporal distribution of the cleavage products of the most abundant AMLEX splice variants including the full length P173, the shorter leucine-rich amelogenin protein (LRAP), and the exon 4-containing P190 in forming enamel, all within the context of the changing enamel matrix proteome during mineralization. We microsampled permanent pig molars, capturing known stages of enamel formation from both crown surface and inner enamel. Nano-LC-MS/MS proteomic analyses after tryptic digestion rendered more than 500 unique protein identifications in enamel, dentin, and bone. We mapped collagens, keratins, and proteolytic enzymes (CTSL, MMP2, MMP10) and determined distributions of P173, LRAP, and P190 products, the enamel proteins enamelin (ENAM) and ameloblastin (AMBN), and matrix-metalloprotease-20 (MMP20) and kallikrein-4 (KLK4). All enamel proteins and KLK4 were near-exclusive to enamel and in excellent agreement with published abundance levels. Phosphorylated P173 and LRAP products decreased in abundance from recently deposited matrix toward older enamel, mirrored by increasing abundances of testicular acid phosphatase (ACPT). Our results showed that hierarchical clustering analysis of secretory enamel links closely matching distributions of unphosphorylated P173 and LRAP products with ACPT and non-traditional amelogenesis proteins, many associated with enamel defects. We report higher protein diversity than previously published and Gene Ontology (GO)-defined protein functions related to the regulation of mineral formation in secretory enamel (e.g., casein α-S1, CSN1S1), immune response in erupted enamel (e.g., peptidoglycan recognition protein, PGRP), and phosphorylation. This study presents a novel approach to characterize and study functional relationships through spatiotemporal mapping of the ephemeral extracellular matrix proteome.

Background and Purpose-Autonomic dysfunction is frequently present in patients with cerebrovascular accidents (CVA).However, the pathophysiological mechanisms of these disorders are not clear. The purpose of the study was to assess the effects of CVA on the autonomic nervous system. Methods-In eight male patients with a history of CVA with damage of the cortical or subcortical structures, we measured the cold pressor response during recording of muscle sympathetic nerve activity (MSNA) from the peroneal nerve on the hemiplegic side. We also studied 10 age-matched male control subjects. Tests were performed before, during, and after immersion of the nonhemiplegic hand in ice water for a period of 3 minutes in each phase. We also recorded changes in heart rate (HR), arterial blood pressure, skin temperature of the middle finger, and perception of pain using the Borg's score. Results-During the control period, the mean burst count of MSNA in CVA (57.2Ϯ3.9 beats/100 HR) was higher than in control subjects (36.3Ϯ3.2 beats/100 HR) (PϽ.05). Total MSNA (the mean burst amplitude per minute times burst rate) increased significantly in CVA and control during the immersion period by 79.9Ϯ18.4% and 133.1Ϯ25.6%, respectively. The percent change in total MSNA in CVA was attenuated during immersion compared with control subjects. The HR and skin temperature responses as well as the Borg's score were similar in both groups during control, hand immersion, and recovery periods. Conclusions-The present results suggest that increased MSNA in CVA may be due to damage of cortical or subcortical structures or stroke-related changes in other areas or nonspecific changes that cause continuous increase in basal MSNA.(Stroke. 1998;29:607-612.)

The study was designed to assess the effects of local heat (LH) application on postganglionic muscle sympathetic nerve activity (MSNA) measured by microneurography in healthy men. In the first protocol, MSNA of the left peroneal nerve, blood pressure (BP), heart rate (HR), and skin temperature of the shin (TSK) were recorded in nine men. In the second protocol, leg blood flow (LBF) was measured in the same subjects by strain-gauge plethysmography. In both protocols, after 10 min of rest in the supine position, a heated hydrocollator pack was applied to the shin and anterior foot for 15 min and recovery was monitored over a period of 20 min. TSK gradually increased from 31.7 ± 0.1 to 41.9 ± 0.5°C (mean ± SEM) during LH. No subject complained of pain, and BP and HR remained constant. The MSNA burst rate (16.1 ± 2.1 beats/min) during the control period decreased significantly (P < 0.05) to 72.0 ± 2.3% during LH. Total MSNA also decreased to 59.2 ± 2.6% (P < 0.05) during LH, but both immediately returned to baseline at recovery. In contrast, LBF in the left leg significantly and immediately increased (P < 0.05) after LH application and remained significantly elevated until the end of the recovery period. These results suggest that: (1) LH application significantly attenuates MSNA without any changes in HR and BP. (2) Other factors in addition to MSNA seem to control regional blood flow in the lower extremity during LH.

The diaphragm of the emphysematous patient is low and limited in its excursions, producing an increased functional residual capacity and decreased pulmonary ventilation. This report describes our experiences with a new technique for 1) the training of abdominal-diaphragmatic (A-D) breathing and 2) the relaxation of accessory respiratory muscles in emphysematous patients. Abdominal muscle contraction during expiration has been shown to increase diaphragmatic excursions and, hence, pulmonary ventilation. Use of this technique has been limited, however, because of the difficulty in learning this breathing pattern. Through continuous audio and visual feedback of myoelectric potentials (myofeedback) from abdominal muscles, 12 patients learned A-D breathing. The lower rectus abdominis muscle was found to be the most suitable muscle for obtaining the myoelectric potentials. Similarly, by providing the patients with myofeedback from their accessory muscles, they decreased the use of these muscles, thus increasing their respiratory efficiency. With myofeedback, patients appear to learn new breathing patterns effectively and in fewer sessions than with conventional procedures.

No abstract