Coated vesicles purified from chick tendon fibroblasts contain newly synthesized type I procollagen

Coated vesicles were purified from embryonic chick tendon fibroblasts pulsed with [3H]proline. They were morphologically and biochemically similar to coated vesicles purified from other sources. Furthermore, they contained newly synthesized Type I procollagen which was protected from bacterial collagenase digestion unless detergent was present. The procollagen remained associated with coated vesicles during immune precipitation and agarose gel electrophoresis. Data from pulse-chase experiments demonstrated that the specific activity of the coated vesicle preparations was approx. 5-fold higher at the 10 min chase point than at either the 0 or 40 min chase points. These data are consistent with the hypothesis that coated vesicles are intermediates in the intracellular transport of newly synthesized Type I procollagen in chick tendon fibroblasts.     

Segregation and linkage analyses of 72 leprosy pedigrees

Data on 72 families with multiple cases of leprosy were analyzed for a susceptibility gene linked to the HLA loci. We conducted segregation analysis with the program POINTER and identity of HLA types by descent analysis to determine the most likely mode of inheritance. We then conducted linkage analysis with the program LINKAS, first assuming linkage equilibrium and then allowing for linkage disequilibrium and etiological heterogeneity. Segregation results suggest a recessive mode of inheritance, especially for the tuberculoid forms of leprosy. The linkage results, limited to tuberculoid forms and assuming a recessive model, suggest a hypothesis of loose linkage with no unlinked locus. When an additive model is assumed, the best fit is obtained with a hypothesis of complete linkage (theta = 0.0) with heterogeneity. We currently favor the additive model as the more plausible one.     

Adaptation of proximal tubular structure and function: insights into compensatory renal hypertrophy

Hypertrophy of the renal tubular cells, especially those of the proximal tubule (PT), accounts for the majority of the increase in kidney size that follows partial removal of renal mass. The propensity of PTs to enlarge appears to be closely linked to an elevation in glomerular filtration rate and may be related to altered tubular fluid flow rate. Hypertrophied PTs reabsorb fluid at an increased rate in vitro, which indicates an intrinsic adaptation of their transport capacity. The hypertrophied cells demonstrate a predominant increase in basolateral membrane area with little change in luminal surface area. This asymmetric structural hypertrophy does not, however, appear to be accompanied by functional asymmetry, for basolateral Na+-K+ pump activity increases roughly in proportion to the increase in cell protein. The activity of the Na+-H+ antiporter, on the other hand, is increased in the brush-border membrane of proximal tubules derived from animals with reduced renal mass. In view of the reported association of Na+-H+ antiport stimulation and mitogenesis in a variety of cell types, the increased activity of this transporter, possibly induced by an increase in tubular fluid flow rate, could be the local stimulus that initiates hypertrophy and determines the organ specificity of the response.     

Internalization, recycling, and redistribution of vasopressin receptors in rat hepatocytes

Three hours after isolation, cultured hepatocytes have approximately 150,000 surface vasopressin receptors/cell, and these exhibit a Kd for 125I-vasopressin of 6 nM based on calculation of Koff/Kon, or a Kd of 9.5 nM based on Scatchard plot analysis. After the binding of 125I-vasopressin to its receptor on the hepatocyte surface, this complex is internalized with a t1/2 of 3-6 min. Following this internalization, the number of vasopressin receptors on the cell surface is restored both in vitro and in the isolated perfused liver with a t1/2 of 8-10 min. This restoration is blocked in vitro by incubation of the hepatocytes at 18 degrees C, but not by cycloheximide, suggesting that internalized vasopressin receptors recycle back to the cell surface. Prolonged incubation of hepatocytes with vasopressin results in the loss of greater than 75% of the vasopressin surface binding at concentrations of vasopressin approximately equivalent to its Kd. The binding of vasopressin to cultured hepatocytes 3-5 h after isolation resembles binding to the isolated perfused whole liver with respect to receptor dynamics. During culture for 48 h, however, we observe a progressive loss of hepatocyte surface vasopressin receptors. Concomitant with this reduction in surface receptors with time in culture, there appears to be a marked elevation in intracellular receptors.     

Calcitonin stimulates cAMP accumulation in chicken osteoclasts

Calcitonin causes an increase in the accumulation of cAMP in mammalian osteoclasts leading to an inhibition of bone resorption. Increases in cAMP subsequent to calcitonin stimulation have not been detected in previous studies of chicken osteoclasts as the only source of large numbers of highly purified cells. In this report, we studied the effects of salmon calcitonin (sCT), bovine parathyroid hormone -(1-34) [(bPTH -(1-34)] and forskolin (FSK) on cAMP accumulation in freshly isolated osteoclasts obtained non-enzymatically from the metaphysis of 5-7 week old rachitogenic chickens. Parathyroid hormone did not stimulate the accumulation of intracellular cAMP. Calcitonin and forskolin treatment caused a nearly 2.5 and 3.5-fold increase in cAMP respectively. This study demonstrates that chicken osteoclasts respond to calcitonin with an increase in cAMP accumulation and that the rachitogenic chicken may be a valuable source of hormonally sensitive cells for the study of osteoclast biology.     

Generic limits re-visited and an updated sectional classification for <Emphasis Type="Italic">Protium</Emphasis> (tribe Protieae). Studies in Neotropical Burseraceae XXV

In recent molecular phylogenetic work on tribe Protieae (Burseraceae) with thorough sampling, the major well-supported clades show remarkable congruence with infrageneric classifications based on morphology, but the results call for synonymizing Crepidospermum and Tetragastris with Protium, thereby making Protieae monophyletic and monogeneric under the name Protium. The necessary synonymies and a brief diagnosis for each of the eight sections of Protium are provided, and these new combinations are made: Protium balsamiferum, P. catuaba, P. chagrense, P. herbertii, P. mucronatum, P. occhionii, and P. rhynchophyllum.     

Key Residues Defining the μ-Opioid Receptor Binding Pocket: A Site-Directed Mutagenesis Study

Abstract: Structural elements of the rat μ-opioid receptor important in ligand receptor binding and selectivity were examined using a site-directed mutagenesis approach. Five single amino acid mutations were made, three that altered conserved residues in the μ, δ, and κ receptors (Asn¹⁵⁰ to Ala, His²⁹⁷ to Ala, and Tyr³²⁶ to Phe) and two designed to test for μ/δ selectivity (Ile¹⁹⁸ to Val and Val²⁰² to Ile). Mutation of His²⁹⁷ in transmembrane domain 6 (TM6) resulted in no detectable binding with [³H]DAMGO (³H-labeled d -Ala², N -Me-Phe⁴,Gly-ol⁵-enkephalin), [³H]bremazocine, or [³H]ethylketocyclazocine. Mutation of Asn¹⁵⁰ in TM3 produces a three- to 20-fold increase in affinity for the opioid agonists morphine, DAMGO, fentanyl, β-endorphin_1–31, JOM-13, deltorphin II, dynorphin_1–13, and U50,488, with no change in the binding of antagonists such as naloxone, naltrexone, naltrindole, and nor-binaltorphamine. In contrast, the Tyr³²⁶ mutation in TM7 resulted in a decreased affinity for a wide spectrum of μ, δ, and κ agonists and antagonists. Altering Val²⁰² to Ile in TM4 produced no change on ligand affinity, but Ile¹⁹⁸ to Val resulted in a four- to fivefold decreased affinity for the μ agonists morphine and DAMGO, with no change in the binding affinities of κ and δ ligands.     

Pulmonary gas exchange is not impaired 24 h after extravehicular activity.

Extravehicular activity (EVA) during spaceflight involves a significant decompression stress. Previous studies have shown an increase in the inhomogeneity of ventilation-perfusion ratio (VA/Q) after some underwater dives, presumably through the embolic effects of venous gas microemboli in the lung. Ground-based chamber studies simulating EVA have shown that venous gas microemboli occur in a large percentage of the subjects undergoing decompression, despite the use of prebreathe protocols to reduce dissolved N(2) in the tissues. We studied eight crewmembers (7 male, 1 female) of the International Space Station who performed 15 EVAs (initial cabin pressure 748 mmHg, final suit pressure either approximately 295 or approximately 220 mmHg depending on the suit used) and who followed the denitrogenation procedures approved for EVA from the International Space Station. The intrabreath VA/Q slope was calculated from the alveolar Po(2) and Pco(2) in a prolonged exhalation maneuver on the day after EVA and compared with measurements made in microgravity on days well separated from the EVA. There were no significant changes in intrabreath VA/Q slope as a result of EVA, although there was a slight increase in metabolic rate and ventilation (approximately 9%) on the day after EVA. Vital capacity and other measures of pulmonary function were largely unaltered by EVA. Because measurements could only be performed on the day after EVA because of logistical constraints, we were unable to determine an acute effect of EVA on VA/Q inequality. The results suggest that current denitrogenation protocols do not result in any major lasting alteration to gas exchange in the lung.