Vascular supply of the anterior interventricular epicardial nerves and ventricular Purkinje fibers in the porcine hearts

The aim of this study was to perform a pilot histological and quantitative analysis of the blood vessels accompanying the epicardial nerves (vasa nervorum) in the porcine hearts. Twenty healthy porcine hearts were used in this study. The blood vessels were analyzed by light microscopy using four different staining techniques in transverse sections taken from the upper, middle, and lower segments of the anterior part of the interventricular region and the adjacent parts of the right and left ventricles containing epicardial nerves and the endocardial peripheral parts of the Purkinje fibers. In total, 317 epicardial nerves were detected. The vasa nervorum were present in 75.7% of these nerves. The vasa nervorum resembled arterioles and postcapillary and collecting venules. One hundred and forty nine epicardial nerves were perivascular, located in the adventitia of the anterior interventricular artery and vein. The remaining 168 nerves ran freely through the epicardial interstitium. The presence of the vasa nervorum was not related to topographical location or nerve diameter. Additionally, from a total of 33 analyzed ventricular complexes of Purkinje fibers small blood vessels located in their proximity were identified in only two cases. It can be concluded that the majority of the anterior epicardial nerves of porcine heart possess well-developed vasa nervorum. In contrast, similar blood vessels are rarely present in the vicinity of the Purkinje fibers. The data obtained contribute to a better understanding of the nutrition of the cardiac nerves.     

Expression of renal cell protein markers is dependent on initial mechanical culture conditions.

The rotating wall vessel is optimized for suspension culture, with laminar flow and adequate nutrient delivery, but minimal shear. However, higher shears may occur in vivo. During rotating wall vessel cultivation of human renal cells, size and density of glass-coated microcarrier beads were changed to modulate initial shear. Renal-specific proteins were assayed after 2 days. Flow cytometry antibody binding analysis of vitamin D receptor demonstrated peak expression at intermediate shears, with 30% reduction outside this range. Activity of cathepsin C showed the inverse pattern, lowest at midshear, with twofold increases at either extreme. Dipeptidyl-peptidase IV had no shear dependence, suggesting that the other results are specific, not universal, changes in membrane trafficking or protein synthesis. On addition of dextran, which changes medium density and viscosity but not shear, vitamin D receptor assay showed no differences from controls. Neither cell cycle, apoptosis/necrosis indexes, nor lactate dehydrogenase release varied between experiments, confirming that the changes are primary, not secondary to cell cycling or membrane damage. This study provides direct evidence that mechanical culture conditions modulate protein expression in suspension culture.     

Survey of Heterorhabditidae and Steinernematidae (Rhabditida,Nematoda) in Western Canada

A survey was done in the summer months along the Alaska Highway, in other parts of British Columbia, in northern Alberta, and in the Yukon Territory for steinernematid and heterorhabditid nematodes occurring in the top 10 cm of soil. Steinernema feltiae and Steinernema spp. were found at 18 and Heterorhabditis megidis at 7 sites of 125 sampled. Most nematodes were found where visible insect infestation occurred and where human influence on the habitat was substantial (e.g., agricultural, forested and bush-hedgerow habitats); none was found in grassland or virgin forests. Heterorhabditis megidis occurred in only the southern, warmer, drier region of British Columbia. In the laboratory some steinernematid isolates and H. megidis killed Galleria mellonella larvae at 13 and 22 C, whereas some isolates of Steinernema killed the larvae at only 13 C. Steinernema spp. from three high altitude sites with low, average July temperatures (13-14 C) are cold-active in that they produced infective juveniles at 13 C and killed G. mellonella at 6 C.     

Sequence regularities and packing of collagen molecules

Fourier analysis of sequences along edges of the type I collagen molecule constructed from two α1(I) and one α2 chains shows that the molecule is two-sided if the supercoil pitch of the α chains along the molecular axis, P, is 39 residues ($\text{D}\text{6}$, where D = 234 residues or 67 nm). One side has alternating charged and hydrophobic regions with spacings of $\text{D}\text{6}$, while the other side has an excess of hydrophobic residues with a spacing of $\text{D}\text{11}$. These characteristics arise from sequence regularities in the α chains and the geometric relationship between the chains. The pattern is marginally strongest with α2 as chain 1. The $\text{D}\text{6}$ sides could form the inside of a helical microfibril where contacts between molecules would fall P apart along the α chains. The $\text{D}\text{11}$ sides could form the outside of the microfibril where contacts between microfibrils would be spaced apart by the α chain supercoil along the microfibril axis, P′. If the microfibril is a 5₄ helix of D-staggered collagen molecules with a left-handed supercoil of pitch $\text{20D}\text{11}$, P′ is close to $\text{2D}\text{11}$ (43 residues). $\text{2D}\text{11}$ subsets in the α chains give rise to the $\text{D}\text{11}$ spacing along the molecule. The microfibril has 4₁ screw symmetry satisfying X-ray diffraction evidence that microfibrils pack in a tetragonal unit cell.This model is the same as proposed previously by us (Trus & Piez, 1976: Piez & Trus, 1977) except that P = 39 rather than 30 residues. Contrary to our earlier assumption, P = 39 residues is within the range allowed by X-ray diffraction measurements. The present results favor P = 39 since it relates regularities in the α chain sequences to helical parameters in a direct way. Furthermore, model studies show that geometric arguments which support P = 30 are equally strong at P = 39 residues.     

Critical assessment of the steady-state Na2SO3 feeding method for kla measurement in fermentors

Important aspects of k(l)a measurement in agitated aerated vessels are briefly characterized from the standpoint of reliability of the measured data. It seems that most of the k(l)a data, based on a number of variants of the steady-state and dynamic methods in noncoalescent liquids, do not have a clear physical meaning, because they are affected by the differences between the actual driving force and the driving force assumed by the model used for its evaluation. A reliability test is given for the Na(2)SO(3) feeding steady-state method (FSM), by comparing the results of air and pure oxygen absorption in a noncoalescent liquid (0.5M Na(2)SO(4) solution) with the results obtained by the independent pressure step dynamic method (RDM). The RDM is one of a few variants of the dynamic method which gives correct k(l)a data unaffected by nonideal mixing of the gas phase in the reactor. It was found that the FSM yields correct k(l)a values only when pure oxygen is used for absorption. When air is absorbed, the FSM gives k(l)a values in the region of k(l)a > 0.1 s(-1) substantially (to 55%) lower than those for pure oxygen absorption.     

Corticotropin-Releasing Hormone Affects Short Immobilization Stress-Induced Changes in Lung Cytosolic and Membrane Glucocorticoid Binding Sites

Glucocorticoids act via glucocorticoid receptors (GR), typically localized in the cytosol (cGR). Rapid action is probably mediated via membrane receptors (mGR). In corticotropin-releasing hormone knockouts (CRH-KO), basal plasma glucocorticoid levels do differ from wild type levels (WT), but are approximately ten times lower during exposure to immobilization stress (IMMO) in comparison to WT. We tested the following hypotheses: (1) the mice lung tissue GR basal numbers would not be changed in CRH-KO (because of similar glucocorticoid levels), (2) the number of GR would be changed in WT but not in KO during short (30, 90, and 120 min) IMMO (because of higher increase of glucocorticoid levels in WT). The basal levels of cGR were not changed in CRH-KO (compared to WT), while mGR were significantly lower (62 %) in CRH-KO. In WT, there was the only decrease (to 32 %) in cGR after 120 min when we also found an increase in mGR in WT (to 201 %). In CRH-KO, IMMO caused gradual decrease in cGR (to 52 % after 30 min, to 46 % after 90 min, and to 32 % after 120 min). In CRH-KO, the only increase in mGR appeared already at 30 min of IMMO. These data suggest, on the contrary to our hypotheses, that CRH-KO are more susceptible to GR changes in early phases of stress.     

Molecular Co-occupancy Identifies Transcription Factor Binding Cooperativity In Vivo

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Synthesis and Some Properties of Modified Oligonucleotides. II. Oligonucleotides Containing 2′-Deoxy-2′-fluoro-β-D-arabinofuranosyl Pyrimidine Nucleosides

Abstract

In order to find the effects of unnatural nucleosides on the stability of duplex, several oligonucleotides containing 1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-uracil(FAU),-cytosine (FAC) and -thymine (FMAU) were synthesized by two alternative approaches: phosphoramidite method on an ABI 392 synthesizer and H-phosphonate procedure on our GeneSyn I universal module synthesizer. It was shown from the melting profiles that the presence of FMAU has a large stabilizing effect on the duplex. Replacement of thymidine with FAU, or deoxycytidine with FAC resulted in the formation of less stable duplexes. Temperature-dependent CD spectroscopy demonstrated that the structures of the fluorine containing oligomers are very similar to those of unmodified oligomers.     

A Bacterial Phosphatase-Like Enzyme of the Malaria Parasite Plasmodium falciparum Possesses Tyrosine Phosphatase Activity and Is Implicated in the Regulation of Band 3 Dynamics during Parasite Invasion

Eukaryotic parasites of the genus Plasmodium cause malaria by invading and developing within host erythrocytes. Here, we demonstrate that PfShelph2, a gene product of Plasmodium falciparum that belongs to the Shewanella-like phosphatase (Shelph) subfamily, selectively hydrolyzes phosphotyrosine, as shown for other previously studied Shelph family members. In the extracellular merozoite stage, PfShelph2 localizes to vesicles that appear to be distinct from those of rhoptry, dense granule, or microneme organelles. During invasion, PfShelph2 is released from these vesicles and exported to the host erythrocyte. In vitro, PfShelph2 shows tyrosine phosphatase activity against the host erythrocyte protein Band 3, which is the most abundant tyrosine-phosphorylated species of the erythrocyte. During P. falciparum invasion, Band 3 undergoes dynamic and rapid clearance from the invasion junction within 1 to 2 s of parasite attachment to the erythrocyte. Release of Pfshelph2 occurs after clearance of Band 3 from the parasite-host cell interface and when the parasite is nearly or completely enclosed in the nascent vacuole. We propose a model in which the phosphatase modifies Band 3 in time to restore its interaction with the cytoskeleton and thus reestablishes the erythrocyte cytoskeletal network at the end of the invasion process.     

Complex Patterns of Chromosome 11 Aberrations in Myeloid Malignancies Target CBL,MLL, DDB1 and LMO2

Exome sequencing of primary tumors identifies complex somatic mutation patterns. Assignment of relevance of individual somatic mutations is difficult and poses the next challenge for interpretation of next generation sequencing data. Here we present an approach how exome sequencing in combination with SNP microarray data may identify targets of chromosomal aberrations in myeloid malignancies. The rationale of this approach is that hotspots of chromosomal aberrations might also harbor point mutations in the target genes of deletions, gains or uniparental disomies (UPDs). Chromosome 11 is a frequent target of lesions in myeloid malignancies. Therefore, we studied chromosome 11 in a total of 813 samples from 773 individual patients with different myeloid malignancies by SNP microarrays and complemented the data with exome sequencing in selected cases exhibiting chromosome 11 defects. We found gains, losses and UPDs of chromosome 11 in 52 of the 813 samples (6.4%). Chromosome 11q UPDs frequently associated with mutations of CBL. In one patient the 11qUPD amplified somatic mutations in both CBL and the DNA repair gene DDB1. A duplication within MLL exon 3 was detected in another patient with 11qUPD. We identified several common deleted regions (CDR) on chromosome 11. One of the CDRs associated with de novo acute myeloid leukemia (P=0.013). One patient with a deletion at the LMO2 locus harbored an additional point mutation on the other allele indicating that LMO2 might be a tumor suppressor frequently targeted by 11p deletions. Our chromosome-centered analysis indicates that chromosome 11 contains a number of tumor suppressor genes and that the role of this chromosome in myeloid malignancies is more complex than previously recognized.