Two human stromal cell lines, HS-5 and HS-27a, represent functionally distinct components of the bone marrow microenvironment.1,2 HS-27a supports cobblestone area formation by early hematopoietic progenitors, whereas HS-5 secretes multiple cytokines that support the proliferation of committed progenitors. These cell lines, which are available from the American Type Culture Collection (ATCC, Manassas, VA), have been distributed to research groups worldwide for use as a tool to understand interactions between hematopoietic cells and their microenvironment. We have recently used DNA microarray technology to characterize and compare the expression of over 17 000 genes in these cell lines.

Briefly, microarray construction and hybridization protocols were modified from Marton et al.3 The microarrays were constructed using a set of more than 17 000 sequence-verified clones from Research Genetics (Huntsville, AL). Of the 17 761 features (spots) on the microarray, 186 are control nonexpressed or nonhuman sequences or housekeeping genes. UniGene cluster IDs could be assigned by GenBank to 16 592 of the features as of June 1, 2001 (UniGene Build 133), indicating that they are representatives of nonredundant unique genes. Many have been functionally characterized, and chromosomal location and tissue expression patterns are known for others. Total RNA was isolated from semiconfluent cultures and reverse-transcribed into cDNA in a nucleotide mix containing amino-allyl deoxyuracil triphosphate (dUTP). The cDNA from stromal cells and Universal RNA (Stratagene, La Jolla, CA) was covalently coupled separately with Cy5 and Cy3 monoreactive fluors, respectively. The Universal RNA consists of a mixture of RNA from 10 different human cell lines with a broad expression coverage of over 80% of the sequences on the array, allowing comparison of expression patterns of multiple different samples of different origin. The Cy5- and Cy3-labeled cDNAs were combined for hybridization to the microarray. Fluorescent array images were collected for both Cy3 and Cy5, and image-intensity data were extracted and analyzed to obtain expression ratios to Universal RNA for each stromal cell line. From these the expression in the 2 lines could be compared.

Here, we present tables identifying the genes with greater than 10-fold and significant differences (Student t test,P < .05) in expression between the 2 cell lines (Tables1 and2). The interleukin-7 receptor, among the genes with much higher expression in HS-27a, is the subject of a separate manuscript appearing in this issue (Iwata et al, page 1318).4 A summary table presenting the complete expression profile for each cell line, as well as a comparison between them, for all 17 000 sequences, is available athttp://parma.fhcrc.org/MIwata. This site also presents technical details of array spotting, hybridization, cDNA synthesis, and fluor-coupling procedures. The entire raw data set, consisting of 4 microarrays per cell line, is publicly available athttp://www.ncbi.nlm.nih.gov/geo/.

Table 1.

Transcripts with greater than 10-fold higher expression in HS-27a than in HS-5

Clone IDAccessionCluster IDNameAnnotationHS-27a/HS-5*
80643 T57803 Hs.10283 RBM8B RNA binding motif protein 8B 83.6 
22355 T89094 Hs.227571 RGS4 Regulator of G-protein signalling 4 77.7 
753982 AA479967 Hs.7882 NA EST 30.7 
429349 AA007419 Hs.227571 RGS4 Regulator of G-protein signalling 4 28.2 
366801 AA029430 Hs.61557 NA EST 27.8 
784337 AA447115 Hs.237356 SDF1 Stromal cell-derived factor 1 25.6 
139354 R63735 Hs.15093 HSPC195 Hypothetical protein 24.4 
841238 AA487121 Hs.237868 IL7R Interleukin 7 receptor 24.1 
68605 T53298 Hs.119206 IGFBP7 Insulin-like growth factor binding protein 7 23.5 
897910 AA598653 Hs.136348 OSF-2 Osteoblast specific factor 2 (fasciclin I-like) 21.0 
489631 AA101875 Hs.81800 CSPG2 Chondroitin sulfate proteoglycan 2 (versican) 19.6 
840460 AA485865 Hs.237868 IL7R Interleukin 7 receptor 19.3 
898218 AA598601 Hs.77326 IGFBP3 Insulin-like growth factor binding protein 3 18.6 
755612 AA419229 Hs.85339 GPR39 G protein-coupled receptor 39 17.2 
773392 AA425749 Hs.2799 CRTL1 Cartilage linking protein 1 16.7  
489519 AA099153 Hs.245188 TIMP3 Tissue inhibitor of metalloproteinase 3 16.4 
302766 N90744 Hs.293907 FLJ23403 Hypothetical protein FLJ23403 15.1 
743146 AA401309 Hs.293907 FLJ23403 Hypothetical protein FLJ23403 13.6 
809719 AA455497 Hs.170121 NA EST 13.4 
133114 R26355 Hs.19545 FZD4 Frizzled (Drosophila) homolog 4 11.7 
1492230 AA875933 Hs.76224 EFEMP1 EGF-containing fibulinlike ECM protein 1 11.6 
796170 AA461086 Hs.16578 NA EST 11.2 
140197 R66101 Hs.103291 LOC51299 Neuritin 10.3 
731311 AA416767 Hs.170121 NA EST 10.3 
309583 N94424 Hs.82547 RARRES1 Retinoic acid receptor responder 10.2 
Clone IDAccessionCluster IDNameAnnotationHS-27a/HS-5*
80643 T57803 Hs.10283 RBM8B RNA binding motif protein 8B 83.6 
22355 T89094 Hs.227571 RGS4 Regulator of G-protein signalling 4 77.7 
753982 AA479967 Hs.7882 NA EST 30.7 
429349 AA007419 Hs.227571 RGS4 Regulator of G-protein signalling 4 28.2 
366801 AA029430 Hs.61557 NA EST 27.8 
784337 AA447115 Hs.237356 SDF1 Stromal cell-derived factor 1 25.6 
139354 R63735 Hs.15093 HSPC195 Hypothetical protein 24.4 
841238 AA487121 Hs.237868 IL7R Interleukin 7 receptor 24.1 
68605 T53298 Hs.119206 IGFBP7 Insulin-like growth factor binding protein 7 23.5 
897910 AA598653 Hs.136348 OSF-2 Osteoblast specific factor 2 (fasciclin I-like) 21.0 
489631 AA101875 Hs.81800 CSPG2 Chondroitin sulfate proteoglycan 2 (versican) 19.6 
840460 AA485865 Hs.237868 IL7R Interleukin 7 receptor 19.3 
898218 AA598601 Hs.77326 IGFBP3 Insulin-like growth factor binding protein 3 18.6 
755612 AA419229 Hs.85339 GPR39 G protein-coupled receptor 39 17.2 
773392 AA425749 Hs.2799 CRTL1 Cartilage linking protein 1 16.7  
489519 AA099153 Hs.245188 TIMP3 Tissue inhibitor of metalloproteinase 3 16.4 
302766 N90744 Hs.293907 FLJ23403 Hypothetical protein FLJ23403 15.1 
743146 AA401309 Hs.293907 FLJ23403 Hypothetical protein FLJ23403 13.6 
809719 AA455497 Hs.170121 NA EST 13.4 
133114 R26355 Hs.19545 FZD4 Frizzled (Drosophila) homolog 4 11.7 
1492230 AA875933 Hs.76224 EFEMP1 EGF-containing fibulinlike ECM protein 1 11.6 
796170 AA461086 Hs.16578 NA EST 11.2 
140197 R66101 Hs.103291 LOC51299 Neuritin 10.3 
731311 AA416767 Hs.170121 NA EST 10.3 
309583 N94424 Hs.82547 RARRES1 Retinoic acid receptor responder 10.2 

Identifiers, names, and functional descriptions are derived from information available in public databases, primarily the National Center for Biotechnology Information (NCBI) UniGene database and GenBank. NA indicates name not available.

EST represents expressed sequence tag; EGF, epidermal growth factor; and ECM, extracellular matrix.

*

Average value in gene expression of HS-27a (n = 4) was divided by average value in gene expression of HS-5 (n = 4). Sequences that appear twice were specified by 2 different clones on the microarray.

Table 2.

Transcripts with greater than 10-fold lower expression in HS-27a than in HS-5

Clone IDAccessionCluster IDNameAnnotationHS-27a/HS5*
845477 AA644211 Hs.196384 PTGS2 Prostaglandin-endoperoxide synthase 2 0.003 
147050 R80217 Hs.196384 PTGS2 Prostaglandin-endoperoxide synthase 2 0.004 
310406 N98591 Hs.93913 IL6 Interleukin 6 0.006 
768417 AA495835 Hs.103839 EPB41L3 Erythrocyte membrane protein band 4.1-like 3 0.010 
324655 W47101 Hs.126256 IL1B Interleukin 1, beta 0.012 
767167 AA424568 Hs.42500 ARL5 ADP-ribosylation factor-like 5 0.017 
503583 AA131240 Hs.297880 NA EST 0.019 
726086 AA399473 Hs.295944 TFPI2 Tissue factor pathway inhibitor 2 0.019 
70692 T49159 Hs.75716 SERPINB2 Serine proteinase inhibitor, clade B, member 2 0.021 
491763 AA150507 Hs.126256 IL1B Interleukin 1, beta 0.021  
323238 W42723 Hs.789 GRO1 GRO1 oncogene 0.023 
773330 AA425450 Hs.82226 GPNMB Glycoprotein (transmembrane) nmb 0.027 
261204 H98218 Hs.308780 NA EST 0.030 
246786 N53172 Hs.23016 RDC1 G protein-coupled receptor 0.031  
46173 H09099 Hs.5378 SPON1 Spondin 1 0.032  
258118 N27108 Hs.43886 NA EST 0.032 
260035 N30372 Hs.54434 IRF5 Interferon regulatory factor 5 0.034  
324437 W46900 Hs.789 GRO1 GRO1 oncogene 0.040  
810859 AA458965 Hs.943 NK4 Natural killer cell transcript 4 0.042 
729924 AA399633 Hs.24872 NA EST 0.045 
814478 AA459263 Hs.227817 BCL2A1 BCL2-related protein A1 0.046  
712049 AA281635 Hs.315463 IL24 Interleukin 24 0.048  
345616 W72431 Hs.82226 GPNMB Glycoprotein (transmembrane) nmb 0.050 
784876 AA448015 Hs.76888 INA Internexin 0.051 
782575 AA447522 Hs.69517 HSJ001348 cDNA for differentially expressed CO16 gene 0.055 
767405 AA417921 Hs.85201 CLECSF2 C-type lectin, superfamily member 2 0.056 
773106 AA425316 Hs.22142 LOC51700 Cytochrome b5 reductase b5R.2 0.060 
181541 H28681 Hs.234074 NA DKFZp761G02121 0.063 
42627 R60995 Hs.21016 COCH Coagulation factor C homology (cochlin) 0.064 
810017 AA455222 Hs.179657 PLAUR Plasminogen activator, urokinase receptor 0.065 
502367 AA134871 Hs.79732 FBLN1 Fibulin 1 0.065 
897768 AA598507 Hs.1640 COL7A1 Collagen, type VII, alpha 1 0.070 
811740 AA463610 Hs.271986 ITGA2 Integrin, alpha 2 (CD49B) 0.071  
285155 N71920 Hs.173560 ODZ2 Odd Oz/ten-m homolog 2 0.075 
129112 R10973 Hs.350197 TSG101 Tumor susceptibility gene 101 protein 0.076 
191882 H38799 Hs.169764 FLJ20701 Hypothetical protein FLJ20701 0.078 
343987 W70234 Hs.44926 DPP4 Dipeptidylpeptidase IV (CD26) 0.078  
357278 W93592 Hs.152213 NA Hypothetical protein FLJ11441 0.079 
162208 H25917 Hs.83583 ARPC2 Actin related protein 2/3 complex, subunit 2 0.079 
469685 AA027856 Hs.7910 RYBP RING1 and YY1 binding protein 0.087  
31825 R41754 Hs.6496 NA EST 0.088 
291880 N67487 Hs.83551 MFAP2 Microfibrillar-associated protein 2 0.090 
280954 N50845 Hs.35089 CNTN3 Contactin 3 0.092 
78148 T61649 Hs.177781 NA Homo sapiens, clone MGC:5618, mRNA 0.094 
324901 W49672 Hs.152213 WNT5A Wingless-type MMTV integration site family 0.095 
241066 H91404 NA NA EST 0.095 
743229 AA400329 Hs.71346 NEF3 Neurofilament 3 0.096 
269815 N27159 Hs.727 INHBA Inhibin, beta A 0.097 
321902 W37448 Hs.348710 NA EST 0.098 
485989 AA040170 Hs.251526 SCYA7 Small inducible cytokine A7 (MCP-3) 0.098 
46055 H08850 Hs.306480 NA DKFZp761E2112 0.099 
840708 AA488084 Hs.177781 NA Homo sapiens, clone MGC:5618, mRNA 0.099 
Clone IDAccessionCluster IDNameAnnotationHS-27a/HS5*
845477 AA644211 Hs.196384 PTGS2 Prostaglandin-endoperoxide synthase 2 0.003 
147050 R80217 Hs.196384 PTGS2 Prostaglandin-endoperoxide synthase 2 0.004 
310406 N98591 Hs.93913 IL6 Interleukin 6 0.006 
768417 AA495835 Hs.103839 EPB41L3 Erythrocyte membrane protein band 4.1-like 3 0.010 
324655 W47101 Hs.126256 IL1B Interleukin 1, beta 0.012 
767167 AA424568 Hs.42500 ARL5 ADP-ribosylation factor-like 5 0.017 
503583 AA131240 Hs.297880 NA EST 0.019 
726086 AA399473 Hs.295944 TFPI2 Tissue factor pathway inhibitor 2 0.019 
70692 T49159 Hs.75716 SERPINB2 Serine proteinase inhibitor, clade B, member 2 0.021 
491763 AA150507 Hs.126256 IL1B Interleukin 1, beta 0.021  
323238 W42723 Hs.789 GRO1 GRO1 oncogene 0.023 
773330 AA425450 Hs.82226 GPNMB Glycoprotein (transmembrane) nmb 0.027 
261204 H98218 Hs.308780 NA EST 0.030 
246786 N53172 Hs.23016 RDC1 G protein-coupled receptor 0.031  
46173 H09099 Hs.5378 SPON1 Spondin 1 0.032  
258118 N27108 Hs.43886 NA EST 0.032 
260035 N30372 Hs.54434 IRF5 Interferon regulatory factor 5 0.034  
324437 W46900 Hs.789 GRO1 GRO1 oncogene 0.040  
810859 AA458965 Hs.943 NK4 Natural killer cell transcript 4 0.042 
729924 AA399633 Hs.24872 NA EST 0.045 
814478 AA459263 Hs.227817 BCL2A1 BCL2-related protein A1 0.046  
712049 AA281635 Hs.315463 IL24 Interleukin 24 0.048  
345616 W72431 Hs.82226 GPNMB Glycoprotein (transmembrane) nmb 0.050 
784876 AA448015 Hs.76888 INA Internexin 0.051 
782575 AA447522 Hs.69517 HSJ001348 cDNA for differentially expressed CO16 gene 0.055 
767405 AA417921 Hs.85201 CLECSF2 C-type lectin, superfamily member 2 0.056 
773106 AA425316 Hs.22142 LOC51700 Cytochrome b5 reductase b5R.2 0.060 
181541 H28681 Hs.234074 NA DKFZp761G02121 0.063 
42627 R60995 Hs.21016 COCH Coagulation factor C homology (cochlin) 0.064 
810017 AA455222 Hs.179657 PLAUR Plasminogen activator, urokinase receptor 0.065 
502367 AA134871 Hs.79732 FBLN1 Fibulin 1 0.065 
897768 AA598507 Hs.1640 COL7A1 Collagen, type VII, alpha 1 0.070 
811740 AA463610 Hs.271986 ITGA2 Integrin, alpha 2 (CD49B) 0.071  
285155 N71920 Hs.173560 ODZ2 Odd Oz/ten-m homolog 2 0.075 
129112 R10973 Hs.350197 TSG101 Tumor susceptibility gene 101 protein 0.076 
191882 H38799 Hs.169764 FLJ20701 Hypothetical protein FLJ20701 0.078 
343987 W70234 Hs.44926 DPP4 Dipeptidylpeptidase IV (CD26) 0.078  
357278 W93592 Hs.152213 NA Hypothetical protein FLJ11441 0.079 
162208 H25917 Hs.83583 ARPC2 Actin related protein 2/3 complex, subunit 2 0.079 
469685 AA027856 Hs.7910 RYBP RING1 and YY1 binding protein 0.087  
31825 R41754 Hs.6496 NA EST 0.088 
291880 N67487 Hs.83551 MFAP2 Microfibrillar-associated protein 2 0.090 
280954 N50845 Hs.35089 CNTN3 Contactin 3 0.092 
78148 T61649 Hs.177781 NA Homo sapiens, clone MGC:5618, mRNA 0.094 
324901 W49672 Hs.152213 WNT5A Wingless-type MMTV integration site family 0.095 
241066 H91404 NA NA EST 0.095 
743229 AA400329 Hs.71346 NEF3 Neurofilament 3 0.096 
269815 N27159 Hs.727 INHBA Inhibin, beta A 0.097 
321902 W37448 Hs.348710 NA EST 0.098 
485989 AA040170 Hs.251526 SCYA7 Small inducible cytokine A7 (MCP-3) 0.098 
46055 H08850 Hs.306480 NA DKFZp761E2112 0.099 
840708 AA488084 Hs.177781 NA Homo sapiens, clone MGC:5618, mRNA 0.099 

Identifiers, names, and functional descriptions are derived from information available in public databases, primarily the National Center for Biotechnology Information (NCBI) UniGene database and GenBank. NA indicates name not available.

*

Average value in gene expression of HS-27a (n = 4) was divided by average value in gene expression of HS-5 (n = 4). Sequences that appear twice were specified by 2 different clones on the microarray.

Gene expression differences in cytokines/chemokines, G-protein signaling molecules, and multiple extracellular matrix proteins add to the known protein and functional characterization of the lines, leading to new insight into the differences in their support function for hematopoietic progenitors.

Supported in part by grants HL62923, CA15704, and DK56465 from the National Institutes of Health, Bethesda, MD.

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