Zhenyu Qin

Phylogeography of the fungal pathogen Histoplasma capsulatum.

Until recently, Histoplasma capsulatum was believed to harbour three varieties, var. capsulatum (chiefly a New World human pathogen), var. duboisii (an African human pathogen) and var. farciminosum (an Old World horse pathogen), which varied in clinical manifestations and geographical distribution. We analysed the phylogenetic relationships of 137 individuals representing the three varieties from six continents using DNA sequence variation in four independent protein‐coding genes. At least eight clades were idengified: (i) North American class 1 clade; (ii) North American class 2 clade; (iii) Latin American group A clade; (iv) Latin American group B clade; (v) Australian clade; (vi) Netherlands (Indonesian?) clade; (vii) Eurasian clade and (viii) African clade. Seven of eight clades represented genetically isolated groups that may be recognized as phylogenetic species. The sole exception was the Eurasian clade which originated from within the Latin American group A clade. The phylogenetic relationships among the clades made a star phylogeny. Histoplasma capsulatum var. capsulatum individuals were found in all eight clades. The African clade included all of the H. capsulatum var. duboisii individuals as well as individuals of the other two varieties. The 13 individuals of var. farciminosum were distributed among three phylogenetic species. These findings suggest that the three varieties of Histoplasma are phylogenetically meaningless. Instead we have to recognize the existence of genetically distinct geographical populations or phylogenetic species. Combining DNA substitution rates of protein‐coding genes with the phylogeny suggests that the radiation of Histoplasma started between 3 and 13 million years ago in Latin America.

The use of THP-1 cells as a model for mimicking the function and regulation of monocytes and macrophages in the vasculature

Since their establishment thirty years ago, THP-1 cells have become one of most widely used cell lines to investigate the function and regulation of monocytes and macrophages in the cardiovascular system. However, because this cell line was derived from the blood of a patient with acute monocytic leukemia, the extent to which THP-1 cells mimic monocytes and macrophages in the vasculature is not entirely known. This article serves as a meaningful attempt to address this question by reviewing the recent publications. The interactions between THP-1 cells and various vascular cells (such as endothelial cells, smooth muscle cells, adipocytes, and T cells) provide insight into the roles of the interconnection of monocytes-macrophages with other vascular cells during vascular inflammation, particularly atherogenesis and obesity. Transcriptome, microRNA profile, and histone modifications of THP-1 cells shed new light on the regulatory mechanism of the monocytes-macrophages in response to various inflammatory mediators, such as oxidized low density lipoprotein, lipopolysaccharide, and glucose. These studies hint that under certain defined conditions, THP-1 cells not only resemble primary monocytes-macrophages isolated from healthy donors or donors with disease, such as diabetes mellitus, but also mimic the in situ alteration of macrophages in the adipose tissue of obese subjects and in atherosclerotic lesions. A potential trajectory is to use this cell line to study the novel molecular mechanisms in monocytes and macrophages in relation to the physiology and pathophysiology of the cardiovascular system, however, the conclusion of studies employing THP-1 cells requires further verification using primary cells and/or in vivo models to be generalized to monocytes and macrophages.

Role of Extracellular Superoxide Dismutase in Hypertension

We previously found that angiotensin II–induced hypertension increases vascular extracellular superoxide dismutase (ecSOD), and proposed that this is a compensatory mechanism that blunts the hypertensive response and preserves endothelium-dependent vasodilatation. To test this hypothesis, we studied ecSOD-deficient mice. ecSOD−/− and C57Blk/6 mice had similar blood pressure at baseline; however, the hypertension caused by angiotensin II was greater in ecSOD−/− compared with wild-type mice (168 versus 147 mm Hg, respectively; P<0.01). In keeping with this, angiotensin II increased superoxide and reduced endothelium-dependent vasodilatation in small mesenteric arterioles to a greater extent in ecSOD−/− than in wild-type mice. In contrast to these findings in resistance vessels, angiotensin II paradoxically improved endothelium-dependent vasodilatation, reduced intracellular and extracellular superoxide, and increased NO production in aortas of ecSOD−/− mice. Whereas aortic expression of endothelial NO synthase, Cu/ZnSOD, and MnSOD were not altered in ecSOD−/− mice, the activity of Cu/ZnSOD was increased by 80% after angiotensin II infusion. This was associated with a concomitant increase in expression of the copper chaperone for Cu/ZnSOD in the aorta but not in the mesenteric arteries. Moreover, the angiotensin II–induced increase in aortic reduced nicotinamide-adenine dinucleotide phosphate oxidase activity was diminished in ecSOD−/− mice as compared with controls. Thus, during angiotensin II infusion, ecSOD reduces hypertension, minimizes vascular superoxide production, and preserves endothelial function in resistance arterioles. We also identified novel compensatory mechanisms involving upregulation of copper chaperone for Cu/ZnSOD, increased Cu/ZnSOD activity, and decreased reduced nicotinamide-adenine dinucleotide phosphate oxidase activity in larger vessels. These compensatory mechanisms preserve large vessel function when ecSOD is absent in hypertension.

Trace metal imaging with high spatial resolution: Applications in biomedicine

New generations of analytical techniques for imaging of metals are pushing hitherto boundaries of spatial resolution and quantitative analysis in biology. Because of this, the application of these imaging techniques described herein to the study of the organization and dynamics of metal cations and metal-containing biomolecules in biological cell and tissue is becoming an important issue in biomedical research. In the current review, three common metal imaging techniques in biomedical research are introduced, including synchrotron X-ray fluorescence (SXRF) microscopy, secondary ion mass spectrometry (SIMS), and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). These are exemplified by a demonstration of the dopamine-Fe complexes, by assessment of boron distribution in a boron neutron capture therapy cell model, by mapping Cu and Zn in human brain cancer and a rat brain tumor model, and by the analysis of metal topography within neuromelanin. These studies have provided solid evidence that demonstrates that the sensitivity, spatial resolution, specificity, and quantification ability of metal imaging techniques is suitable and highly desirable for biomedical research. Moreover, these novel studies on the nanometre scale (e.g., of individual single cells or cell organelles) will lead to a better understanding of metal processes in cells and tissues.

Essential role for the Menkes ATPase in activation of extracellular superoxide dismutase: Implication for vascular oxidative stress

Extracellular superoxide dismutase (SOD3), a secretory copper enzyme, plays an important role in atherosclerosis and hypertension by modulating the levels of extracellular superoxide anion (O2 •–) in the vasculature. Little is known about the mechanisms by which SOD3 obtains its catalytic copper cofactor. Menkes ATPase (MNK) has been shown to transport cytosolic copper to the secretory pathway in nonvascular cells. We performed the present study to determine whether MNK is required for the activation of SOD3 in the vasculature. Here we show that MNK was highly expressed in the various vascular tissues and cells. Aortas and cultured fibroblasts from MNK mutant (MNKmut) mice showed a marked decrease in specific activity of SOD3, but not SOD1 (cytosolic form), which was partially restored by copper addition. Copper treatment in wild‐type cells promoted the direct interaction and colocalization of SOD3 with MNK in the trans‐Golgi network (TGN), suggesting that MNK transports copper to SOD3 in the TGN. Aortas of MNKmut mice revealed a decrease in activity of SOD3, but not SOD1, in association with a robust increase in O2 •– levels. Finally, both MNK and SOD3 proteins were highly expressed in the intimal lesions of atherosclerotic vessels. In conclusion, vascular MNK plays an essential role in full activity of SOD3 through transporting copper to SOD3 in the TGN, thereby regulating O2 •– levels in the vasculature. These studies provide a novel insight into vascular MNK as a critical modulator of “superoxide” stress, which may contribute to cardiovascular disease.

Bioimaging of Metals and Biomolecules in Mouse Heart by Laser Ablation Inductively Coupled Plasma Mass Spectrometry and Secondary Ion Mass Spectrometry

Bioimaging mass spectrometric techniques allow direct mapping of metal and biomolecule distributions with high spatial resolution in biological tissue. In this study laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS) was used for imaging of transition metals (Fe, Cu, Zn, Mn, and Ti), alkali and alkaline-earth metals (Na, K, Mg, and Ca, respectively), and selected nonmetals (such as C, P, and S) in native cryosections of mouse heart. The metal and nonmetal images clearly illustrated the shape and the anatomy of the samples. Zinc and copper were inhomogeneously distributed with average concentrations of 26 and 11 μg g(-1), respectively. Titanium and manganese were detected at concentrations reaching 1 and 2 μg g(-1), respectively. The highest regional metal concentration of 360 μg g(-1)was observed for iron in blood present in the lumen of the aorta. Secondary ion mass spectrometry (SIMS) as an elemental and biomolecular mass spectrometric technique was employed for imaging of Na, K, and selected biomolecules (e.g., phosphocholine, choline, cholesterol) in adjacent sections. Here, two different bioimaging techniques, LA-ICPMS and SIMS, were combined for the first time, yielding novel information on both elemental and biomolecular distributions.

Extracellular superoxide dismutase (ecSOD) in vascular biology: an update on exogenous gene transfer and endogenous regulators of ecSOD

Extracellular superoxide dismutase (ecSOD) is the major extracellular scavenger of superoxide (O(2)(.-)) and a main regulator of nitric oxide (NO) bioactivity in the blood vessel wall, heart, lungs, kidney, and placenta. Involvement of O(2)(.-) has been implicated in many pathological processes, and removal of extracellular O(2)(.-) by ecSOD gene transfer has emerged as a promising experimental technique to treat vascular disorders associated with increased oxidant stress. In addition, recent studies have clarified mechanisms that regulate ecSOD expression, tissue binding, and activity, and they have provided new insight into how ecSOD interacts with other factors that regulate vascular function. Finally, studies of a common gene variant in humans associated with disruption of ecSOD tissue binding suggest that displacement of the enzyme from the blood vessel wall may contribute to vascular diseases. The purpose of this review is to summarize recent research findings related to ecSOD function and gene transfer and to stimulate other investigations into the role of this unique antioxidant enzyme in vascular pathophysiology and therapeutics.

Role of Menkes ATPase in Angiotensin II-Induced Hypertension: A Key Modulator for Extracellular Superoxide Dismutase Function

The extracellular superoxide dismutase (SOD3), a secretory copper-containing enzyme, regulates angiotensin II (Ang II)–induced hypertension by modulating levels of extracellular superoxide anion. The present study was designed to determine the role of the copper transporter Menkes ATPase (MNK) in Ang II–induced SOD3 activity and hypertension in vivo. Here we show that chronic Ang II infusion enhanced systolic blood pressure and vascular superoxide anion production in MNK mutant (MNKmut) mice as compared with those in wild-type mice, which are associated with impaired acetylcholine-induced endothelium-dependent vasorelaxation in MNKmut mice. These effects in MNKmut mice are rescued by infusion of the SOD mimetic Tempol. By contrast, norepinephrine-induced hypertension, which is not associated with an increase in vascular superoxide anion production, is not affected in MNKmut mice. Mechanistically, basal and Ang II infusion-induced increase in vascular SOD3-specific activity is significantly inhibited in MNKmut mice. Coimmunoprecipitation analysis reveals that Ang II stimulation promotes association of MNK with SOD3 in cultured vascular smooth muscle cell and in mouse aortas, which may contribute to SOD3-specific activity by increasing copper delivery to SOD3 through MNK. In summary, MNK plays an important role in modulating Ang II–induced hypertension and endothelial function by regulating SOD3 activity and vascular superoxide anion production and becomes a potential therapeutic target for oxidant stress-dependent cardiovascular diseases.

CD14 directs adventitial macrophage precursor recruitment: role in early abdominal aortic aneurysm formation.

Background Recruitment of macrophage precursors to the adventitia plays a key role in the pathogenesis of abdominal aortic aneurysms (AAAs), but molecular mechanisms remain undefined. The innate immune signaling molecule CD14 was reported to be upregulated in adventitial macrophages in a murine model of AAA and in monocytes cocultured with aortic adventitial fibroblasts (AoAf) in vitro, concurrent with increased interleukin‐6 (IL‐6) expression. We hypothesized that CD14 plays a crucial role in adventitial macrophage precursor recruitment early during AAA formation. Methods and Results CD14−/− mice were resistant to AAA formation induced by 2 different AAA induction models: aortic elastase infusion and systemic angiotensin II (AngII) infusion. CD14 gene deletion led to reduced aortic macrophage infiltration and diminished elastin degradation. Adventitial monocyte binding to AngII‐infused aorta in vitro was dependent on CD14, and incubation of human acute monocytic leukemia cell line‐1 (THP‐1) monocytes with IL‐6 or conditioned medium from perivascular adipose tissue (PVAT) upregulated CD14 expression. Conditioned medium from AoAf and PVAT induced CD14‐dependent monocyte chemotaxis, which was potentiated by IL‐6. CD14 expression in aorta and plasma CD14 levels were increased in AAA patients compared with controls. Conclusions These findings link CD14 innate immune signaling via a novel IL‐6 amplification loop to adventitial macrophage precursor recruitment in the pathogenesis of AAA.

Transepithelial Transport Characteristics of the Antihypertensive Peptide, Lys-Val-Leu-Pro-Val-Pro, in Human Intestinal Caco-2 Cell Monolayers

An antihypertensive peptide, Lys-Val-Leu-Pro-Val-Pro (KVLPVP), can reduce blood pressure in hypertensive rats after being orally administered. In this study, the transepithelial transport of intact KVLPVP was examined by Caco-2 monolayers. The results were as follows: (i) The flux was not saturable for apical (AP) to basolateral (BL) or BL-AP transport when the concentration of KVLPVP was 1–8 mM. (ii) Sodium deoxycholate loosened the tight junction in the Caco-2 cells and significantly improved the transport process. (iii) Phenylarsine oxide, a transcytotic process inhibitor, had little effect on the transport process. (iv) The influx and eflux of KVLPVP remained unchanged in the presence of the ATP inhibitor sodium azide. (v) This transport was not inhibited by the peptide transporter substrates Gly-Pro or arphanine A. All these data indicate that paracellular transport diffusion was the major flux mechanism for the intact KVLPVP.