Transport of Intranasally Instilled Fine Fe2O3 Particles into the Brain: Micro-distribution, Chemical States, and Histopathological Observation

It has been demonstrated that inhaled fine (d < 2.5 μm) and ultrafine (d < 100 nm) particles produce more severe toxicity than coarse particles. Some recent data support the concept that the central nervous system (CNS) may be a target for the inhaled fine particulates. This work describes initial observation of the transport of intranasally instilled fine ferric oxide (Fe₂O₃) particles in animal brain. The iron micro-distribution and chemical state in the mice olfactory bulb and brain stem on day 14 after intranasal instillation of fine Fe₂O₃ particle (280 ± 80 nm) suspension at a single dose of 40 mg/kg body weight were analyzed by synchrotron radiation x-ray fluorescence and x-ray absorption near-edge structure (XANES). The micro-distribution map of iron in the olfactory bulb and brain stem shows an obvious increase of Fe contents in the olfactory nerve and the trigeminus of brain stem, suggesting that Fe₂O₃ particles were possibly transported via uptake by sensory nerve endings of the olfactory nerve and trigeminus. The XANES results indicate that the ratios of Fe (III)/Fe (II) were increased in the olfactory bulb and brain stem. The further histopathological observation showed that the neuron fatty degeneration occurred in the CA3 area of hippocampus. Such results imply an adverse impact of inhalation of fine Fe₂O₃ particles on CNS.     

Probing the role of microenvironment for microencapsulated Sacchromyces cerevisiae under osmotic stress

Cell encapsulation opens a new avenue to the oral delivery of genetically engineered microorganism for therapeutic purpose. Osmotic stress is one of the universal chemical stress factors in the application of microencapsulation technology. In order to understand the effect and mechanism of the encapsulated microenvironment on protecting cells from hyper-osmotic stress, yeast cells of Saccharomyces cerevisiae Y800 were encapsulated in calcium alginate micro-gel beads (MB), alginate-chitosan-alginate (ACA) solid core microcapsules (SCM), and ACA liquid core microcapsules (LCM), respectively. The stress-induced intracellular components and enzyme activity including trehalose, glycerol and super oxide dismutase (SOD) were measured. Free cell culture was used as control. The survival of encapsulated cells and the cells released from MB, SCM and LCM after osmotic shock induced by NaCl solution (1, 2 and 3M) was evaluated. An analysis method was established to probe the effect of encapsulated microenvironment on the cell tolerance to osmotic stress. The results showed that LCM gave rise to the highest level of intracellular trehalose and glycerol, and SOD activity, as well as the highest survival rate of encapsulated cells or cells released from microcapsule. It was demonstrated that LCM was able to induce the highest stress response and stress tolerance of cells, which was adapted during culture, while SCM failed. The theoretical analysis revealed that it was the liquid alginate matrix in microcapsule that played a central role in domesticating the cells to adapt to hyper-osmotic stress. This finding provides a very useful guideline to cell encapsulation.     

MicroRNA regulation and interspecific variation of gene expression

MicroRNAs (miRNAs) modulate expression of their target genes in various tissues and at different developmental stages, but it is unclear whether they drive cross-species variation in gene expression. By comparing data from mammal and fly species we found that the cross-species expression variation of miRNA targets is significantly lower than that of other genes. This implies that miRNAs can affect gene expression by reducing stochastic noise, buffering cross-species variation and constraining evolutionary gene expression variation.     

Cloning and characterization of a plastidic glycerol 3-phosphate dehydrogenase cDNA from Dunaliella salina

A cDNA encoding a nicotinamide adenine dinucleotide (NAD+) -dependent glycerol 3-phosphate dehydrogenase (GPDH) has been cloned by rapid amplification of cDNA ends from Dunaliella salina. The cDNA is 3032 base pairs long with an open reading frame encoding a polypeptide of 701 amino acids. The polypeptide shows high homology with published NAD+ -dependent GPDHs and has at its N-terminal a chloroplast targeting sequence. RNA gel blot analysis was performed to study GPDH gene expression under different conditions, and changes of the glycerol content were monitored. The results indicate that the cDNA may encode an osmoregulated isoform primarily involved in glycerol synthesis. The 701-amino-acid polypeptide is about 300 amino acids longer than previously reported plant NAD+ -dependent GPDHs. This 300-amino-acid fragment has a phosphoserine phosphatase domain. We suggest that the phosphoserine phosphatase domain functions as glycerol 3-phosphatase and that, consequently, NAD+ -dependent GPDH from D. salina can catalyze the step from dihydroxyacetone phosphate to glycerol directly. This is unique and a possible explanation for the fast glycerol synthesis found in D. salina.     

Receptor-mediated and bulk-phase endocytosis cause macrophage and cholesterol accumulation in Niemann-Pick C disease

These studies explored the roles of receptor-mediated and bulk-phase endocytosis as well as macrophage infiltration in the accumulation of cholesterol in the mouse with Niemann-Pick type C (NPC) disease. Uptake of LDL-cholesterol varied from 514 microg/day in the liver to zero in the central nervous system. In animals lacking LDL receptors, liver uptake remained about the same (411 microg/day), but more cholesterol was taken up in extrahepatic organs. This uptake was unaffected by the reductive methylation of LDL and consistent with bulk-phase endocytosis. All tissues accumulated cholesterol in mice lacking NPC1 function, but this accumulation was decreased in adrenal, unchanged in liver, and increased in organs like spleen and lung when LDL receptor function was also deleted. Over 56 days, the spleen and lung accumulated amounts of cholesterol greater than predicted, and these organs were heavily infiltrated with macrophages. This accumulation of both cholesterol and macrophages was increased by deleting LDL receptor function. These observations indicate that both receptor-mediated and bulk-phase endocytosis of lipoproteins, as well as macrophage infiltration, contribute to the cholesterol accumulation seen in NPC disease. These macrophages may also play a role in parenchymal cell death in this syndrome.     

Regulation of TRPV5 Single-Channel Activity by Intracellular pH

The transient receptor potential channel TRPV5 contributes to the apical entry pathway for transcellular calcium reabsorption in the kidney. Acid load causes hypercalciuria in animals and humans. We have previously reported that intracellular protons directly inhibit TRPV5. Here, we examined the effects of intracellular pH on single-channel activity of TRPV5. We found that TRPV5 channels exhibit full and subconductance open states in excised inside–out patches of Chinese hamster ovary cells. The slope conductance values (Na⁺ as a charge carrier, between −25 and −75 mV) for full and subconductance opening at intracellular pH 7.4 were 59 ± 6 and 29 ± 3 pS, respectively. Intracellular acidification caused a small decrease in single-channel conductance. Importantly, intracellular acidification decreased open probability for the full and subconductance states and increased probability for closing. To investigate how intracellular protons decrease open probability of the channel, we proposed a simple three-state model for open–subconductance–closed state transition and examined the effects of acidification on the respective forward and reverse rate constants. We found that intracellular acidification decreases opening of TRPV5 predominantly by promoting a transition from the subconductance to the closed state. Thus, intracellular acidification directly inhibits TRPV5 by causing a conformational change(s) leading to a decrease of open probability of TRPV5 as well as of the single-channel conductance. Seung-Kuy Cha and Wasey Jabbar contributed equally to this work.     

Substance P is associated with heart enlargement and apoptosis in murine dilated cardiomyopathy induced by Taenia crassiceps infection

Dilated cardiomyopathy (degeneration of heart muscle and heart enlargement) is an important cause of heart failure among young adults. Dilated cardiomyopathy may be a complication during or after various viral, bacterial, or parasitic diseases. Substance P (SP) is a neurotransmitter that is involved in the pathogenesis of various diseases. To determine whether SP is associated with cardiac changes in murine cysticercosis, we compared heart-weight to body-weight ratio, cardiac pathology, cardiomyocyte size, and cardiac-apoptosis (TUNEL assay) in hearts from Taenia crassiceps-infected (wild-type vs. SP-knockout) mice. We noted that, as compared with control uninfected wild-type mice, elevated protein levels of SP and its receptor as studied by ELISA or immunohistochemistry, respectively, were elevated in the hearts of parasite-infected wild-type mice. The heart-weight to body-weight ratios were significantly higher in the parasite-infected wild-type mice versus those of the infected SP-knockout mice. Furthermore, wild-type infected mice developed dilated cardiomyopathy with increased chamber size of both ventricles, decreased ventricular wall thickness, compensatory cardiomyocyte hypertrophy, and increased cardiac apoptosis. This cardiac pathology did not develop in mice lacking SP activity (i.e., in infected SP knockout mice) or in uninfected mice. These data indicate that SP is associated with cardiac changes in an animal model of parasitic dilated cardiomyopathy.