HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) A corrigendum has been published All Versions of this Article: 12/3/209    most recent 00081.2001v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (16) Citing Articles via Google Scholar Google Scholar Articles by Hoshikawa, Y. Articles by Geraci, M. W. Search for Related Content PubMed PubMed Citation Articles by Hoshikawa, Y. Articles by Geraci, M. W.

Hypoxia induces different genes in the lungs of rats compared with mice

¹ Pulmonary Hypertension Center
² Division of Pulmonary Sciences and Critical Care Medicine
³ Cardiovascular Pulmonary Research Laboratory
⁴ Department of Pathology
⁵ Department of Pharmacology, University of Colorado Health Sciences Center, Denver, Colorado 80262
⁶ Department of Thoracic Surgery, Institute of Development, Aging, and Cancer, Tohoku University, Sendai, Japan 980-8575

	ABSTRACT

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

 
Different animal species have a varying response to hypoxia. Mice develop less pulmonary artery thickening after chronic hypoxia exposure than rats. We hypothesized that the lung tissue gene expression pattern displayed in hypoxic rats would differ from that of hypoxic mice. We exposed Sprague-Dawley rats and C57BL/6 mice to both 1 and 3 wk of hypobaric hypoxia. Although both species developed pulmonary hypertension, mice showed less pulmonary vascular remodeling than rats. Microarray gene analysis demonstrated a distinct pattern of gene expression between mice and rats when exposed to hypoxic conditions. In addition, some genes appeared to be more responsive at an earlier time point of 1 wk of hypoxia. Hypoxic conditions in the rat induce genes involved in endothelial cell proliferation, repression of apoptosis, and vasodilation. Mice exposed to hypoxic conditions decrease the expression of genes involved in vasodilation and in endothelial cell proliferation. Although we cannot determine whether the differential expression of genes during chronic hypoxia is cause or consequence of the differential pulmonary vascular remodeling, we propose that a balance between over- and under-expression of a selective group of genes may be responsible for lung vascular remodeling and vascular tone control.

pulmonary hypertension; chronic hypoxia; pulmonary vascular remodeling; gene microarray analysis; species differences

	INTRODUCTION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

 
CHRONIC HYPOXIA CAUSES pulmonary hypertension (PH) in humans (3) and in most animals (17, 18, 32, 36, 40, 43, 44). However, the degree of the structural alteration of the lung vessels (pulmonary vascular remodeling) varies between species (18, 43). For example, hamsters are known to develop less muscularization of their precapillary arteries during chronic hypoxia than rats (43), and coati mundi have already very muscular precapillary arteries at sea level and do not develop further vascular structural alterations when exposed to chronic hypoxia (20, 21) . It therefore appears that the extent of the lung vascular remodeling in response to chronic hypoxia and chronic hypoxic vasoconstriction is genetically determined.

Lung vascular remodeling associated with chronic hypoxia has been recently investigated in transgenic mice, demonstrating that chronic PH and lung vascular remodeling is decreased in mice with a knockout of the 5-lipoxygenase gene (42) and also in mice which overexpress the gene encoding the critical enzyme prostacyclin synthase in surfactant protein-synthesizing lung cells (13). In contrast, mice with targeted disruption of the gene encoding the prostacyclin receptor develop a greater degree of lung vascular remodeling during chronic hypoxia compared with the wild type (24).

For the same hypoxic challenge, the pulmonary vascular remodeling is less pronounced in mice compared with rats (23, 24). Even within a species, the extent of vascular remodeling is variable and strain specific, with both the BALB/c (19) and FVB/N mice (13) demonstrating remodeling, whereas the C57BL/6 strain shows minimal changes in medial wall thickness when exposed to chronic hypoxia (12). Therefore, we examined the alterations in the lung tissue gene expression pattern in Sprague-Dawley (SD) rats and in C57BL/6 mice kept at Denver altitude (DA) or exposed to hypobaric hypoxia at both 1 and 3 wk time points. Our microarray gene expression data show that there are categorically different gene expression responses triggered by exposure to chronic hypoxia in rat and in mouse lungs. The balance between upregulation of the genes encoding for cell proliferation and downregulation of genes encoding for vascular tone may be associated with the greater degree of vascular remodeling and higher pulmonary artery pressures in the rat.

	METHODS

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

 
Hypobaric Hypoxic Exposure
For exposure to hypobaric hypoxia, 8-wk-old male C57BL/6 mice (n = 3) and 7-wk-old male SD rats (n = 3) were housed in a hypobaric chamber for 1 and 3 wk at a simulated altitude of 17,000 ft. (Pb = 410 mmHg; inspired PO₂ = 76 mmHg). Three mice and three rats were kept at DA (5,280 ft.; Pb = 630 mmHg, inspired PO₂ = 122 mmHg) for more than 3 wk. Evaluation at 1 wk of hypoxia was included to determine whether gene expression was altered at an earlier time point and to investigate the resultant changes in pulmonary artery pressure.

Measurement of Right Ventricular Pressure of Mice and Pulmonary Artery Pressure of Rats
Hemodynamic assessment was performed as previously described by direct measurement of right ventricular systolic pressure (RVSP) for mice (12, 13) and by measurement of mean pulmonary artery pressure (mPAP) utilizing a catheter inserted through the jugular vein for rats (23, 37). After RVSP or mPAP measurement, lung tissue was inflated with low-melt agarose and fixed with 10% buffered formalin.

Details for mice.
Briefly, the mice were anesthetized with ketamine and xylazine (100 mg/kg and 15 mg/kg, respectively) by way of a hindquarter intramuscular injection. After adequate sedation, the animals were placed in a supine position, breathing room air. The pressure transducer was calibrated to a zero point at the mid-anteroposterior diameter of the chest. A 26-gauge needle was introduced percutaneously into the thorax through a subxyphoid approach. Right ventricular pressure was measured using a pressure transducer (Gulton-Statham, Costa Mesa, CA) and recorded on a multichannel recorder (Grass Instruments, Quincy, MA). Approximately 10 measurements per animal were made, and the average pressure value was recorded.

Details for rats.
The rats were anesthetized with ketamine and xylazine (60 mg/kg and 8 mg/kg, respectively) intramuscularly. The pulmonary arterial pressure was measured after a polyethylene catheter (PE-10; Intramedic, Clay Adams, Parsippany, NJ) had been inserted into the main pulmonary artery through the right jugular vein. The shape of the pressure tracing displayed on an oscilloscope guided the placement of the catheter. The pressure was measured using a pressure transducer (Gulton-Statham); the zero reference point was the level of the midchest. Approximately 10 measurements per animal were made, and the average pressure value was recorded.

Histology and Morphometric Analysis
A representative hematoxylin and eosin-stained lung section through the hilum was coded and blindly evaluated for remodeling of pulmonary arteries in the range of 30–50 µm for mice and 10–150 µm for rats as previously described (13, 23, 39). Pulmonary arteries were differentiated from pulmonary veins by the identification of the double elastic lamina in the vascular walls of the arteries using phase contrast. The wall thickness of each artery (VWT) was expressed as percentage of the vessel diameter by the formula VWT = (Ed - Id)/Ed x 100, where Ed is external diameter and Id is internal diameter. Between 10 and 20 measurements were made per animal (3 animals for each condition), and the average was calculated.

Gene Microarray Analysis
Three C57BL/6 mice and three SD rats were exposed to hypobaric hypoxia for 1 and 3 wk. Three mice and three rats were kept at DA. The animals were euthanized by lethal injection of sodium pentobarbital (Abbott Laboratories, North Chicago, IL). Lungs were excised, frozen in liquid nitrogen, and stored at -70°C. Total RNA was extracted from the lung tissues by using the RNeasy Total RNA Isolation Kit (Qiagen, Valencia, CA). Fifteen micrograms of total RNA was collected from each sample and evaluated individually. Twenty micrograms of the labeled cRNA mixture was applied to the GeneChip microarray analysis (Affymetrix, Santa Clara, CA) as described previously (14, 15, 34). Rat samples were hybridized to the Affymetrix Rat Genome U34A arrays, whereas murine samples were hybridized to the Murine Genome Array U74Av2 for these studies, unless otherwise stated. Detailed protocols for data analysis of Affymetrix microarrays and extensive documentation of the sensitivity and quantitative aspects of the method have been described (16, 28, 31)

Quantitative RT-PCR Analysis for the Rat Osteopontin Gene
To corroborate findings by microarray analysis, quantitative RT-PCR was performed on the gene encoding for osteopontin as described previously (15). The primers used were forward 5'-CAAGGCCTCTGGGAAGCA-3' and reverse 5'-CGATTCCGCAGCTCAGTTC-3' (63-bp band product). For the internal control, we quantitated ribosomal RNA content in each template using TaqMan ribosomal RNA control reagents (Perkin-Elmer) and normalized the copy number of each gene.

Statistical Analysis
Analysis of gene expression was conducted by previously reported Affymetrix software. All values were expressed as means ± SE. Means of several groups were compared by three methods including unpaired t-test, analysis of variance (ANOVA), and a matrix analysis of all possible combinations of baseline (DA) to each of the hypoxic conditions (1 wk and 3 wk). In this matrix analysis, a very stringent requirement was that all nine of nine possible comparisons (a 3 x 3 table created for each condition) must show concordance in the "increase" or "decrease" call generated by the Affymetrix Microarray Suite 5.0 (MAS 5.0) algorithm.

Prior to analysis by any of the statistical tests, variability filters were applied to the dataset to minimize multiple testing errors. The first filter uses the Affymetrix mRNA detection "call" (MAS version 5.0) to exclude all genes with an "absolute call" of "absent" in all experiments. The second filter identifies genes with at least moderate variance over the entire experiment; genes that do not vary at all cannot possibly be related to the system under study, which in this case is the hypoxia exposure. Since we can assume that most genes are not related to the system under study (hypoxia exposure), then the gene with the median variance is a reasonable model of null variation, that is, the variation due to other factors. We calculate the variances for each gene. The null hypothesis is that these variances represent random and normally distributed noise. We can then compute the statistic W = (N - 1) s2/median (s2), where N is the number of observations of the gene, which is approximately chi-square distributed with N - 1 degree of freedom (22), and s2 is the variance of each gene, and median (s2) is the median over all the gene variances. We calculate a P value for rejecting the null hypothesis that the gene did not vary, and perform the "false discovery rate" multiple testing correction (5), setting the false discovery rate to be 10%. This results in a list of genes with significantly greater variation than the median variation gene, with at most 10% of that list including genes having true variation less than or equal to median variation. These preprocessing steps screen out genes with low variance and low levels of mRNA expression. The remainder of the genes is then fed to the Kruskal-Wallis or ANOVA test and FDR correction (10% in this case), producing a final list of genes whose expression levels are significantly different between at least two time points.

Two-tailed parametric analysis was considered significant when P < 0.01. Analysis by the three separate methods produced multiple genes that were altered in the setting of hypoxia. A subset of genes that was significantly altered by two of the three statistical methods is reported. In all cases, every attempt was made to compare the same genes that were statistically different between the two species. This was accomplished by utilizing the NetAFFX website (http://www.affymetrix.com/analysis/index.affx) to cross reference the genes represented on the different arrays. If there was representation on both the rat and the murine arrays, this is noted in the tables. If the genes were statistically changed in one species, but the corresponding gene was not represented on the array from the other species, then it is noted in the tables; a symbol "" represents genes which are not represented on the array, and therefore their expression is undetermined.

	RESULTS

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

 
Both Rats and Mice Develop Pulmonary Hypertension After Exposure to Hypobaric Hypoxia
Both 1 and 3 wk of hypobaric hypoxia caused a significant increase in mPAP in SD rats. After 1 wk of hypoxia, PAP values rose from 18.3 ± 0.3 to 24.8 ± 2.8. mmHg (P < 0.01). At 3 wk of hypoxia, pressures rose even higher compared with control [n = 3 vs. control, n = 3; 33.3 ± 1.3 vs. 18.3 ± 0.3 mmHg; P < 0.001 (Fig. 1A)]. C57BL/6 mice displayed higher RVSP after exposure to 3 wk of hypobaric hypoxia compared with control animals kept at DA (28.0 ± 1.0 mmHg at DA vs. 35.6 ± 1.5 at 3 wk, P = 0.3 x 10^-3). Although there was a trend toward higher RVSP after the 1 wk exposure, this did not reach statistical significance [control, n = 3; 28.0 ± 1.0 mmHg at DA; 30.7 ± 0.5, at 1 wk; P = not significant (NS)] (Fig. 1B).

View larger version (21K): [in this window] [in a new window]   Fig. 1. Mean pulmonary artery pressure (mPAP) of rats (A) and right ventricular systolic pressure (RVSP) of C57BL/6 mice (B) at Denver altitude (DA) and after exposure to hypobaric hypoxia at both 1 and 3 wk. A: after 1 wk of hypoxia, PAP values rose from 18.3 ± 0.3 to 24.8 ± 2.8 mmHg (P < 0.01). At 3 wk of hypoxia, pressures rose even higher compared with control (n = 3 vs. control, n = 3; 33.3 ± 1.3 vs. 18.3 ± 0.3 mmHg; *P < 0.001). B: C57BL/6 mice displayed higher RVSP after exposure to 3 wk of hypobaric hypoxia compared with control animals kept at DA (28.0 ± 1.0 at DA vs. 35.6 ± 1.5 mmHg at 3 wk; **P = 0.3 x 10-3). Although there was a trend toward higher RVSP after the 1 wk exposure, this did not reach statistical significance (control, n = 3; 28.0 ± 1.0 at DA vs. 30.7 ± 0.5 mmHg at 1 wk; P = NS).

View larger version (77K): [in this window] [in a new window]   Fig. 2. A: percent vessel wall thickness of small- to medium-sized (10–150 µm for rats and 30–50 µm for mice) pulmonary arteries at DA and after chronic exposure to hypobaric hypoxia. Percent vessel wall thickness was calculated by external diameter (Ed) minus internal diameter (Id) divided by the Ed multiplied by 100: (Ed - Id)/Ed x 100. Hypoxic rats (n = 3) developed significantly more severe vessel wall thickening than control rats (n = 3) (39.5 vs. 29.9%; *P < 0.0001), whereas control (n = 10) and hypoxic (n = 13) mice had similar vessel wall thickness (16.3 vs. 16.3%; NS). B: histology of the pulmonary arteries from a rat kept at DA (a), a rat exposed to hypobaric hypoxia for 3 wk (b), a mouse kept at DA (c), and a chronically hypoxic mouse (d). Note the marked increase in the pulmonary vessel wall thickness of the chronically hypoxic rat, whereas the hypoxic mouse did not develop any vessel wall thickening.

View larger version (11K): [in this window] [in a new window]   Fig. 3. Genes upregulated in the hypoxic rat lungs but unchanged by chronic hypoxia in C57BL/6 mouse lungs. The names of each of the 9 genes and the details of the ratio changes are given in Table 1. The genes are shown in relative expression changes, for each individual animal.

View larger version (12K): [in this window] [in a new window]   Fig. 4. Genes decreased in expression for the hypoxic rat lungs but unchanged in the hypoxic mouse lungs. The names of each of the 19 genes and the details of the ratio changes are given in Table 2. The genes are shown in relative expression changes, for each individual animal.

View larger version (11K): [in this window] [in a new window]   Fig. 5. Genes upregulated in expression for the hypoxic mouse lungs but unchanged in the rat lungs. The names of each of the 5 genes and the details of the ratio changes are given in Table 3. The genes are shown in relative expression changes, for each individual animal.

View larger version (10K): [in this window] [in a new window]   Fig. 6. Genes downregulated in expression for the hypoxic mouse lungs but unchanged in the rat lungs. The names of each of the 7 genes and the details of the ratio changes are given in Table 4. The genes are shown in relative expression changes, for each individual animal.

Quantitative RT-PCR of the Osteopontin Gene in Rat Lungs
The osteopontin gene in the hypoxic rat lungs had a 7.4-fold increased expression compared with that in the rat lungs kept at DA. When we assessed the lung osteopontin gene expression using quantitative RT-PCR, we confirmed the microarray expression data (Fig. 7).

View larger version (13K): [in this window] [in a new window]   Fig. 7. Quantitative RT-PCR of the osteopontin mRNA in the lungs from rats kept at DA (n = 3) and exposed to hypoxia for 3 wk (n = 3). Dots indicate relative expression of osteopontin gene normalized by ribosomal RNA expression in each rat lung; horizontal bars indicate average value in each group of rats. The chronically hypoxic rat lungs showed a 14- to 92-fold higher expression of the osteopontin gene than did DA control rat lungs.

	DISCUSSION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

We expected that the stimulus of chronic hypoxia would result in an altered lung tissue gene expression pattern when compared with the control lung tissue, since hypoxia-inducible factor 1 (HIF-1) and egr-1 have been shown to be involved in the transcriptional regulation of a large number of genes (33, 46). Surprisingly, a relatively small number of genes were altered in their expression in the lungs after both 1 and 3 wk of hypoxia. Our novel approach of lung tissue gene expression profiling makes use of the absent lung vascular remodeling in the mouse and compares lung tissue gene expression patterns in chronically hypoxic mice and rats which do show a robust pulmonary arterial muscularization response. Based on our gene expression data, new specific hypotheses can be developed and strategies to test causal relationships.

Although descriptive, our study expands the spectrum of factors that may need consideration and functional analysis. Previous work by a number of investigators has focused on the role of extracellular matrix proteins (7), matrix metalloproteinases (35), mediators of inflammation (38), and growth factors (41), which all very likely contribute to the process of vascular remodeling in chronically hypoxic lungs. We have recently used the gene microarray technique to analyze lungs from patients with primary PH (PPH) (14) and lungs from prostacyclin receptor (PGI-R) knockout (KO) and wild-type mice exposed to chronic hypobaric hypoxia. We see the tissue gene expression data as a first step of exploration, as a survey of candidate genes that are frequently "syn"-expressed in a pattern that may suggest a mechanism. In this context, it is of interest that chronic hypoxia affects C57BL/6 mice and SD rats differently and that the gene expression pattern in the lungs from patients with PPH, which carry plexiform lesions composed of proliferating endothelial cells (38), is very different from that in pulmonary hypertensive rat lungs where plexiform lesions do not form. It is also of interest that genes varied in their time response to hypoxia. Some exhibited significant alterations at 1 wk, whereas others exhibited alterations at 3 wk.

Integrin-, secretory phospholipase A₂, CXC chemokine, and phosphoinositide 3-kinase, which all have roles in proliferation and remodeling, were significantly elevated in the rat but unchanged in the mouse. In addition, osteopontin, which may have a role in reducing vascular smooth muscle cell apoptosis, was also elevated. In contrast, mice had less upregulation of genes responsible for proliferation and remodeling and demonstrated a downregulation in genes encoding for proteins responsible for vasodilation. It has been reported that vascular smooth muscle cells of the synthetic phenotype but not of the contractile phenotype express osteopontin mRNA (45) and that osteopontin reduces vascular smooth muscle cell apoptosis.

The phosphoinositide 3-kinase gene had upregulated expression in the hypoxia-exposed rat lungs but not in the mouse lungs. This kinase has been shown in vascular smooth muscle cells to regulate the expression of p27KIP1 leading to an enhanced G₁/S transition of vascular smooth muscle cells (4); p21 also plays a major role in IGF-I-stimulated migration and proliferation of vascular smooth muscle cells. The PP2A gene was upregulated in the hypoxia-exposed rat lungs but not in the mouse lungs. PP2A is a phosphatase with a role in inflammation, lipid mediator production, and proliferation of smooth muscle cells (2). Integrin- was significantly upregulated in the hypoxic rat lungs but not in the mouse lungs. Authors have recognized that the class of integrins along with endothelins may play a role in remodeling via proliferation, migration, and apoptosis of smooth muscle cells (30).

Mice exposed to hypoxic conditions exhibited a very different pattern of gene expression that favored downregulation of genes responsible for vascular smooth muscle proliferation and vasodilation. In contrast, upregulated genes in the hypoxic rat lungs were unchanged in the mouse. Downregulated genes included IGF binding protein, leptin receptor, and TGF-ß binding protein. In animal models, IGF has been recognized as likely having a role in stimulating smooth muscle hyperplasia (8).

Leptin is an endocrine hormone that has recently received a great deal of attention in the field of research in obesity. Studies in leptin-deficient mice suggest that leptin has a role in inducing vascular permeability and stimulating angiogenesis in adipose tissue (6). Furthermore, cellular studies have concluded that hypoxia may regulate pro-angiogenic properties of leptin (1). Whether leptin is regulated by hypoxic conditions in other vascular beds is unclear but likely warrants further investigation. TGF-ß binding protein appears to have a role in rats in smooth muscle cell migration and subsequent arteriole intimal thickening (26).

Data from our microarray experiments can be compared with previously published work examining gene expression changes during exposure to hypoxia. One surprising finding of our gene profiling study is that neither the genes encoding tenascin nor the serotonin transporter were upregulated in their expression in the hypoxic rats, in light of recent publications which assigned an important role for this matrix protein (25) and the serotonin transporter in hypoxia-induced pulmonary vascular remodeling in the rat (9–11). We have previously demonstrated that overexpression of the prostacyclin synthase gene in transgenic mice confers complete protection from the development of PH (13). Furthermore, patients with PH have decreased expression of the prostacyclin synthase gene (37). In the current expression analysis, prostacyclin synthase was elevated only in rats and was significant at the 1-wk time point. Other investigators have demonstrated that the endothelin receptors are important and involved in pulmonary vascular remodeling during chronic hypoxia. Li et al. (29) described the increased level of expression of endothelin and ET-A as well as ET-B receptors in rats exposed to chronic hypoxia. We could not detect a statistical difference in gene expression using our stringent criteria for change. Le Cras et al. (27) have investigated the regulation of iNOS and eNOS in chronic hypoxia in the rat lung and have demonstrated modest increases (1.4- to 2.1-fold change). The expressions of these transcripts are below the level of detection and are called "absent" using our technology. These findings underscore some of the differences between our approach and previous detection strategies. In the lung, there are more than 40 distinct cell types that contribute to the transcriptome of a whole lung preparation. Determining the specific cell type responsible for the overexpression would require further analysis using different techniques, such as in situ hybridization. Western analysis and immunohistochemistry could be used for the evaluation of protein expression.

In summary, the identical stimulus, chronic hypoxia, produces a very different gene expression pattern in rat lungs when compared with mouse lungs. Whether this difference is the cause or the consequence of the lack of hypoxia-induced pulmonary vascular remodeling in the mouse should be a focus of future investigation. In addition, since there were only two time points examined (1 and 3 wk), it is difficult to determine the timing of sequential genetic events. We compared our microarray results to those of other investigators who examined hypoxic response. In some instances, there were discrepancies that can be explained by several factors. First, genes were considered statistically significant if at least two of three statistical tests indicated significance, and, as a result, some genes of interest were only significant by one method. Secondly, some genes of interest were not present on our arrays. This underscores some of the present limitations of this technology. However, we propose that a group of genes, rather than a single gene, is critically involved in pulmonary vascular remodeling and also that the genes encoding proteins involved in vascular tone regulation are likely different from those involved in lung vessel cell growth.

	ACKNOWLEDGMENTS

 
We acknowledge the University of Colorado Comprehensive Cancer Center Gene Expression Core Facility for the assistance with the microarray experiments.

This work has been supported by National Heart, Lung, and Blood Institute Grants HL-43180 (to N. F. Voelkel and M. W. Geraci) and HL-66254 (to R. M. Tuder, N. F. Voelkel, and M. W. Geraci) and by the Shirley K. Wisham Memorial Research Foundation for Pulmonary Hypertension Research.

	FOOTNOTES

 
Article published online before print. See web site for date of publication (http://physiolgenomics.physiology.org).

Address for reprint requests and other correspondence: M. W. Geraci, Division of Pulmonary Sciences and Critical Care Medicine, Univ. of Colorado Health Sciences Center Campus Box C-272, 4200 E. Ninth Ave., Denver, CO 80262 (E-mail: mark.geraci{at}uchsc.edu).

10.1152/physiolgenomics.00081.2001.

^* Y. Hoshikawa and P. Nana-Sinkam contributed equally to this work.

	References

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION References

 

1. Ambrosini G, Nath AK, Sierra-Honigmann MR, and Flores-Riveros J. Transcriptional activation of the human leptin gene in response to hypoxia. Involvement of hypoxia-inducible factor 1. J Biol Chem 277: 34601–34609, 2002.[Abstract/Free Full Text]
2. Antonio V, Brouillet A, Janvier B, Monne C, Bereziat G, Andreani M, and Raymondjean M. Transcriptional regulation of the rat type IIA phospholipase A2 gene by cAMP and IL-1ß in vascular smooth muscle cells: interplay of C/EBP, NF-B, and Ets transcription factors. Biochem J 368: 415–424, 2002.[Medline]
3. Arias-Stella J and Saldana M. Terminal portion of the pulmonary arterial in people native to high altitudes. Circulation 28: 915–925, 1963.[Free Full Text]
4. Bacqueville D, Casagrande F, Perret B, Chap H, Darbon JM, and Breton-Douillon M. Phosphatidylinositol 3-kinase inhibitors block aortic smooth muscle cell proliferation in mid-late G1 phase: effect on cyclin-dependent kinase 2 and the inhibitory protein p27KIP1. Biochem Biophys Res Commun 244: 630–636, 1998.[Web of Science][Medline]
5. Benjamini Y and Hochberg Y. Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc B57: 289–300, 1995.
6. Cao R, Brakenhielm E, Wahlestedt C, Thyberg J, and Cao Y. Leptin induces vascular permeability and synergistically stimulates angiogenesis with FGF-2 and VEGF. Proc Natl Acad Sci USA 98: 6390–6395, 2001.[Abstract/Free Full Text]
7. Cowan KN, Jones PL, and Rabinovitch M. Elastase and matrix metalloproteinase inhibitors induce regression, and tenascin-C antisense prevents progression, of vascular disease. J Clin Invest 105: 21–34, 2000.[Web of Science][Medline]
8. Dempsey EC, Stenmark KR, McMurtry IF, O’Brien RF, Voelkel NF, and Badesch DB. Insulin-like growth factor I and protein kinase C activation stimulate pulmonary artery smooth muscle cell proliferation through separate but synergistic pathways. J Cell Physiol 144: 159–165, 1990.[Web of Science][Medline]
9. Eddahibi S, Hanoun N, Lanfumey L, Lesch KP, Raffestin B, Hamon M, and Adnot S. Attenuated hypoxic pulmonary hypertension in mice lacking the 5-hydroxytryptamine transporter gene. J Clin Invest 105: 1555–1562, 2000.[Web of Science][Medline]
10. Eddahibi S, Humbert M, Fadel E, Raffestin B, Darmon M, Capron F, Simonneau G, Dartevelle P, Hamon M, and Adnot S. Serotonin transporter overexpression is responsible for pulmonary artery smooth muscle hyperplasia in primary pulmonary hypertension. J Clin Invest 108: 1141–1150, 2001.[Web of Science][Medline]
11. Eddahibi S, Raffestin B, Hamon M, and Adnot S. Is the serotonin transporter involved in the pathogenesis of pulmonary hypertension? J Lab Clin Med 139: 194–201, 2002.[Web of Science][Medline]
12. Fagan KA, Fouty BW, Tyler RC, Morris KG Jr, Hepler LK, Sato K, Le Cras TD, Abman SH, Weinberger HD, Huang PL, McMurtry IF, and Rodman DM. The pulmonary circulation of homozygous or heterozygous eNOS-null mice is hyperresponsive to mild hypoxia. J Clin Invest 103: 291–299, 1999.[Web of Science][Medline]
13. Geraci MW, Gao B, Shepherd DC, Moore MD, Westcott JY, Fagan KA, Alger LA, Tuder RM, and Voelkel NF. Pulmonary prostacyclin synthase overexpression in transgenic mice protects against development of hypoxic pulmonary hypertension. J Clin Invest 103: 1509–1515, 1999.[Web of Science][Medline]
14. Geraci MW, Moore M, Gesell T, Yeager ME, Alger L, Golpon H, Gao B, Loyd JE, Tuder RM, and Voelkel NF. Gene expression patterns in the lungs of patients with primary pulmonary hypertension: a gene microarray analysis. Circ Res 88: 555–562, 2001.[Abstract/Free Full Text]
15. Golpon HA, Geraci MW, Moore MD, Miller HL, Miller GJ, Tuder RM, and Voelkel NF. HOX genes in human lung: altered expression in primary pulmonary hypertension and emphysema. Am J Pathol 158: 955–966, 2001.[Abstract/Free Full Text]
16. Golub TR, Slonim DK, Tamayo P, Huard C, Gaasenbeek M, Mesirov JP, Coller H, Loh ML, Downing JR, Caligiuri MA, Bloomfield CD, and Lander ES. Molecular classification of cancer: class discovery and class prediction by gene expression monitoring. Science 286: 531–537, 1999.[Abstract/Free Full Text]
17. Grover RF, Reeves JT, Weir EK, McMurtry IF, and Alexander AF. Genetic transmission of susceptibility to hypoxic pulmonary hypertension. Prog Respir Res 9: 112–117. 1975.
18. Grover RF, Vogel JK, Averill KH, and Blount SG. Pulmonary hypertension: individual and species variability relative to vascular reactivity. Am Heart J 66: 1–3, 1963.[Web of Science][Medline]
19. Hales CA, Kradin RL, Brandstetter RD, and Zhu YJ. Impairment of hypoxic pulmonary artery remodeling by heparin in mice. Am Rev Respir Dis 128: 747–751, 1983.[Web of Science][Medline]
20. Hanson WL, Boggs DF, Kay JM, Hofmeister SE, Okada O, and Wagner WW Jr. Pulmonary vascular response of the coati to chronic hypoxia. J Appl Physiol 88: 981–986, 2000.[Abstract/Free Full Text]
21. Hanson WL, Boggs DF, Kay JM, Hofmeister SE, and Wagner WW Jr. Collateral ventilation and pulmonary arterial smooth muscle in the coati. J Appl Physiol 74: 2219–2224, 1993.[Abstract/Free Full Text]
22. Hogg RV and Craig AT. Introduction to Mathematical Statistics. Upper Saddle River, NJ: Prentice Hall, 1995.
23. Hoshikawa Y, Ono S, Suzuki S, Tanita T, Chida M, Song C, Noda M, Tabata T, Voelkel NF, and Fujimura S. Generation of oxidative stress contributes to the development of pulmonary hypertension induced by hypoxia. J Appl Physiol 90: 1299–1306, 2001.[Abstract/Free Full Text]
24. Hoshikawa Y, Voelkel NF, Gesell TL, Moore MD, Morris KG, Alger LA, Narumiya S, and Geraci MW. Prostacyclin receptor-dependent modulation of pulmonary vascular remodeling. Am J Respir Crit Care Med 164: 314–318, 2001.[Abstract/Free Full Text]
25. Jones PL, Chapados R, Baldwin HS, Raff GW, Vitvitsky EV, Spray TL, and Gaynor JW. Altered hemodynamics controls matrix metalloproteinase activity and tenascin-C expression in neonatal pig lung. Am J Physiol Lung Cell Mol Physiol 282: L26–L35, 2002.[Abstract/Free Full Text]
26. Kanzaki T, Shiina R, Saito Y, Oohashi H, and Morisaki N. Role of latent TGF-ß1 binding protein in vascular remodeling. Biochem Biophys Res Commun 246: 26–30, 1998.[Web of Science][Medline]
27. Le Cras TD, Xue C, Rengasamy A, and Johns RA. Chronic hypoxia upregulates endothelial and inducible NO synthase gene and protein expression in rat lung. Am J Physiol Lung Cell Mol Physiol 270: L164–L170, 1996.[Abstract/Free Full Text]
28. Lee CK, Klopp RG, Weindruch R, and Prolla TA. Gene expression profile of aging and its retardation by caloric restriction. Science 285: 1390–1393, 1999.[Abstract/Free Full Text]
29. Li H, Chen SJ, Chen YF, Meng QC, Durand J, Oparil S, and Elton TS. Enhanced endothelin-1 and endothelin receptor gene expression in chronic hypoxia. J Appl Physiol 77: 1451–1459, 1994.[Abstract/Free Full Text]
30. Liu G, Eskin SG, and Mikos AG. Integrin vß3 is involved in stimulated migration of vascular adventitial fibroblasts by basic fibroblast growth factor but not platelet-derived growth factor. J Cell Biochem 83: 129–135, 2001.[Medline]
31. Lockhart DJ, Dong H, Byrne MC, Follettie MT, Gallo MV, Chee MS, Mittmann M, Wang C, Kobayashi M, Horton H, and Brown EL. Expression monitoring by hybridization to high-density oligonucleotide arrays. Nat Biotechnol 14: 1675–1680, 1996.[Web of Science][Medline]
32. Rabinovitch M, Gamble W, Nadas AS, Miettinen OS, and Reid L. Rat pulmonary circulation after chronic hypoxia: hemodynamic and structural features. Am J Physiol Heart Circ Physiol 236: H818–H827, 1979.[Abstract/Free Full Text]
33. Semenza GL. Surviving ischemia: adaptive responses mediated by hypoxia-inducible factor 1. J Clin Invest 106: 809–812, 2000.[Web of Science][Medline]
34. Sugita M, Geraci M, Gao B, Powell RL, Hirsch FR, Johnson G, Lapadat R, Gabrielson E, Bremnes R, Bunn PA, and Franklin WA. Combined use of oligonucleotide and tissue microarrays identifies cancer/testis antigens as biomarkers in lung carcinoma. Cancer Res 62: 3971–3979, 2002.[Abstract/Free Full Text]
35. Thompson K and Rabinovitch M. Exogenous leukocyte and endogenous elastases can mediate mitogenic activity in pulmonary artery smooth muscle cells by release of extracellular-matrix bound basic fibroblast growth factor. J Cell Physiol 166: 495–505, 1996.[Web of Science][Medline]
36. Tucker A, McMurtry IF, Reeves JT, Alexander AF, Will DH, and Grover RF. Lung vascular smooth muscle as a determinant of pulmonary hypertension at high altitude. Am J Physiol 228: 762–767, 1975.[Abstract/Free Full Text]
37. Tuder RM, Cool CD, Geraci MW, Wang J, Abman SH, Wright L, Badesch D, and Voelkel NF. Prostacyclin synthase expression is decreased in lungs from patients with severe pulmonary hypertension. Am J Respir Crit Care Med 159: 1925–1932, 1999.[Abstract/Free Full Text]
38. Tuder RM, Groves B, Badesch DB, and Voelkel NF. Exuberant endothelial cell growth and elements of inflammation are present in plexiform lesions of pulmonary hypertension. Am J Pathol 144: 275–285, 1994.[Abstract]
39. Voelkel NF, Allard JD, Anderson SM, and Burke TJ. cGMP and cAMP cause pulmonary vasoconstriction in the presence of hemolysate. J Appl Physiol 86: 1715–1720, 1999.[Abstract/Free Full Text]
40. Voelkel NF and Tuder RM. Hypoxia-induced pulmonary vascular remodeling: a model for what human disease? J Clin Invest 106: 733–738, 2000.[Web of Science][Medline]
41. Voelkel NF, Tuder RM, Bridges J, and Arend WP. Interleukin-1 receptor antagonist treatment reduces pulmonary hypertension generated in rats by monocrotaline. Am J Respir Cell Mol Biol 11: 664–675, 1994.[Abstract]
42. Voelkel NF, Tuder RM, Wade K, Hoper M, Lepley RA, Goulet JL, Koller BH, and Fitzpatrick F. Inhibition of 5-lipoxygenase-activating protein (FLAP) reduces pulmonary vascular reactivity and pulmonary hypertension in hypoxic rats. J Clin Invest 97: 2491–2498, 1996.[Web of Science][Medline]
43. Walker BR, Berend N, and Voelkel NF. Comparison of muscular pulmonary arteries in low and high altitude hamsters and rats. Respir Physiol 56: 45–50, 1984.[Medline]
44. Will DH, Alexader AF, Reeves JT, and Grover RF. High altitude-induced pulmonary hypertension in normal cattle. Circ Res 10: 172–177, 1962.[Abstract/Free Full Text]
45. Yamamoto M, Aoyagi M, Azuma H, and Yamamoto K. Changes in osteopontin mRNA expression during phenotypic transition of rabbit arterial smooth muscle cells. Histochem Cell Biol 107: 279–287, 1997.[Web of Science][Medline]
46. Yan SF, Fujita T, Lu J, Okada K, Shan ZY, Mackman N, Pinsky DJ, and Stern DM. Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress. Nat Med 6: 1355–1361, 2000.[Web of Science][Medline]

This article has been cited by other articles:

				E. Daley, C. Emson, C. Guignabert, R. de Waal Malefyt, J. Louten, V. P. Kurup, C. Hogaboam, L. Taraseviciene-Stewart, N. F. Voelkel, M. Rabinovitch, et al. Pulmonary arterial remodeling induced by a Th2 immune response J. Exp. Med., February 18, 2008; 205(2): 361 - 372. [Abstract] [Full Text] [PDF]

				T. M. Bull, C. D. Coldren, M. W. Geraci, and N. F. Voelkel Gene Expression Profiling in Pulmonary Hypertension Proceedings of the ATS, January 1, 2007; 4(1): 117 - 120. [Abstract] [Full Text] [PDF]

				T. Weng, Z. Chen, N. Jin, L. Gao, and L. Liu Gene expression profiling identifies regulatory pathways involved in the late stage of rat fetal lung development Am J Physiol Lung Cell Mol Physiol, November 1, 2006; 291(5): L1027 - L1037. [Abstract] [Full Text] [PDF]

				M. D Ganfornina, M. T Perez-Garcia, G Gutierrez, E Miguel-Velado, J. R Lopez-Lopez, A Marin, D Sanchez, and C Gonzalez Comparative gene expression profile of mouse carotid body and adrenal medulla under physiological hypoxia J. Physiol., July 15, 2005; 566(2): 491 - 503. [Abstract] [Full Text] [PDF]

				S. A. Gharib, D. L. Luchtel, D. K. Madtes, and R. W. Glenny Global gene annotation analysis and transcriptional profiling identify key biological modules in hypoxic pulmonary hypertension Physiol Genomics, June 16, 2005; 22(1): 14 - 23. [Abstract] [Full Text] [PDF]

				P. Li, S. Oparil, W. Feng, and Y.-F. Chen Hypoxia-responsive growth factors upregulate periostin and osteopontin expression via distinct signaling pathways in rat pulmonary arterial smooth muscle cells J Appl Physiol, October 1, 2004; 97(4): 1550 - 1558. [Abstract] [Full Text] [PDF]

				R. L. Keith, Y. E. Miller, T. M. Hudish, C. E. Girod, S. Sotto-Santiago, W. A. Franklin, R. A. Nemenoff, T. H. March, S. P. Nana-Sinkam, and M. W. Geraci Pulmonary Prostacyclin Synthase Overexpression Chemoprevents Tobacco Smoke Lung Carcinogenesis in Mice Cancer Res., August 15, 2004; 64(16): 5897 - 5904. [Abstract] [Full Text] [PDF]

				M. R. MacLean, G. A. Deuchar, M. N. Hicks, I. Morecroft, S. Shen, J. Sheward, J. Colston, L. Loughlin, M. Nilsen, Y. Dempsie, et al. Overexpression of the 5-Hydroxytryptamine Transporter Gene: Effect on Pulmonary Hemodynamics and Hypoxia-Induced Pulmonary Hypertension Circulation, May 4, 2004; 109(17): 2150 - 2155. [Abstract] [Full Text] [PDF]

				N.F. Voelkel and C.D. Cool Pulmonary vascular involvement in chronic obstructive pulmonary disease Eur. Respir. J., November 2, 2003; 22(46_suppl): 28s - 32s. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) A corrigendum has been published All Versions of this Article: 12/3/209    most recent 00081.2001v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (16) Citing Articles via Google Scholar Google Scholar Articles by Hoshikawa, Y. Articles by Geraci, M. W. Search for Related Content PubMed PubMed Citation Articles by Hoshikawa, Y. Articles by Geraci, M. W.