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Gregory J Essayan


A novel gene, designated ML-1, was identified from a human genomic DNA clone and human T cell cDNA sequences. The second exon of ML-1 gene shares significant sequence identity with the gene encoding IL-17 (IL-17). ML-1 gene expression was up-regulated in activated PBMCs, CD4+ T cells, allergen-specific Th0, Th1, and Th2 clones, activated basophils, and mast cells. Increased expression of the ML-1 gene, but not IL-17, was seen following allergen challenge in four asthmatic subjects, suggesting its role in allergic inflammatory responses. ML-1 from transiently transfected COS-7 cells was able to induce gene expression and protein production for IL-6 and IL-8 (at 10 ng/ml of ML-1: for IL-6, 599.6 ± 19.1 pg/ml; for IL-8, 1724.2 ± 132.9 pg/ml; and at 100 ng/ml of ML-1: for IL-6, 1005.3 ± 55.6 pg/ml; for IL-8, 4371.4 ± 280.5 pg/ml; p < 0.05 for both doses vs baseline) in primary bronchial epithelial (PBE) cells. Furthermore, increased expression of ICAM-1 was found in ML-1-stimulated PBE cells (mean fluorescence intensity (MFI) = 31.42 ± 4.39 vs baseline, MFI = 12.26 ± 1.77, p < 0.05), a functional feature distinct from IL-17 (MFI = 11.07 ± 1.22). This effect was not inhibited by a saturating amount of IL-17. These findings demonstrate that ML-1 is a novel cytokine with a distinct function, and suggest a different receptor for ML-1 on PBE cells.

The maintenance of normal physiology and the expression of various inflammatory diseases involve highly interactive molecular and cellular networks. Dysregulation of the cytokine network is an important contributing factor in allergic inflammatory diseases, such as asthma. Understanding of this intricate network, along with characterization of relevant novel genes, would aid in discovering the molecular basis of inflammatory diseases. Recent advances in human genome sequencing and in the use of bioinformatics tools have facilitated the process for gene discovery and identifying its functional correlate, particularly for those genes with cell type-specific expression patterns. The newly discovered gene can be integrated in the search for the molecular basis of various diseases.

Recent cloning and sequencing studies have demonstrated a family of IL-17-related genes with potential proinflammatory functions. Human IL-17 is a T cell-derived, homodimeric protein and exhibits pleiotropic biological activities (1, 2, 3). IL-17 is able to stimulate the production of IL-6, IL-8, G-CSF, stem cell factor, and PGE2 from various cell types, such as fibroblasts, keratinocytes, and renal epithelial cells (3, 4, 5, 6, 7). Although the expression of IL-17 is restricted to activated T cells, the IL-17R is widely expressed. In addition, elevated IL-17 mRNA expression has been found in mononuclear cells from patients with multiple sclerosis (8), in patients with rheumatoid arthritis (9), and in patients with systemic lupus erythematosus (10), suggesting a role for IL-17 in the initiation or maintenance of inflammatory responses. In the airway, IL-17 induces expression of the C-X-C chemokines, IL-8 and macrophage inflammatory protein 2, which selectively recruit neutrophils into the airway (11). Moreover, IL-17 synergizes, but not by itself, with IFN-γ in the induction of ICAM-1 expression on epithelial cells (12); the induction of ICAM-1 is associated with airway inflammation seen in bronchial asthma (13).

As part of our ongoing molecular genetic studies of complex diseases, a potential coding region sequence with sequence homology with IL-17 was initially identified from a human genomic DNA clone by the use of bioinformatics tools and sequencing. We describe herein functional characterization of this novel cytokine gene and demonstrate its expression in an inflammatory disease.

Materials and Methods

Full-length cDNA sequence of the ML-1 gene

To obtain a full-length cDNA sequence, both 5′- and 3′-RACE were performed using cDNAs from activated ragweed allergen-specific T cells as templates. For 3′-RACE, cDNAs were amplified using poly(dT) and a predicted exon sequence primer, 5′-GGCATCATCAATGAAAACCAG-3′. The PCR products were run on 1% low-melting agarose gel, purified using a GeneClean kit (Qbiogene, Carlsbad, CA), and subjected to a nested PCR using an internal sequence primer, 5′-TTCCATGTCACGTAACATCG-3′. After PCR, the products were cloned and sequenced. For 5′-RACE, cDNAs were first tailed with poly(dA) oligonucleotides using TdT enzyme, purified using Sephadex G25 spin columns, and subjected to nested PCRs using poly(dT), a coding region sequence (5′-TCACCAGCACCTTCTCCAAC-3′) and an internal sequence (5′-AAGAAACAGAGCAGCCTTGG-3′) primer. The PCR products were then cloned and sequenced.

Analysis of tissue distribution, RNA isolation, and cellular expression of ML-1

Tissue distribution data for IL-17 and ML-1 were acquired using Rapid-Scan gene expression panels for human tissues (OriGene Technologies, Rockville, MD) according to the manufacturer’s instructions with 5 mM magnesium and ML-1- and IL-17-specific primer pairs. The sequences of primers for ML-1 were as follows: forward, 5′-GGCATCATCAATGAAAACCAG-3′; reverse, 5′-TCACCAGCACCTTCTCCAAC-3′. The sequences of primers for IL-17 were: forward, 5′-ACTCCTGGGAAGACCTCATTG-3′; reverse, 5′-GGCCACATGGTGGACAATCG-3′. PCR products were visualized by ethidium bromide-containing gel and photographed. Tissues were graded on a 0–4 grading system based on visualization of bands at the concentrations of cDNA provided by the manufacturer (++++ = product obtained with >1 pg/ml; +++ = product obtained with >10 pg/ml; ++ = product obtained with >100 pg/ml; + = product obtained with 1000 pg/ml; grade 0 = no product obtained with 1000 pg/ml). Appropriate normalization of cDNA provided by the manufacturer was confirmed by PCR amplification for the constitutive marker gene, β-actin.

PBMCs were isolated from the blood of allergic subjects. Peripheral blood CD4+, CD8+ T cells, and monocytes were isolated (>90% purity) from a normal healthy individual using RosetteSep kits from StemCell Technology (Vancouver, Canada) according to the manufacturer’s instruction. Human allergen-specific T cell clones were generated by limiting dilution cloning and subcloning from two atopic subjects, followed by biweekly stimulation of T cells with ragweed allergen extract or purified Amb a 1 (a major ragweed allergen) along with irradiated autologous PBMCs as APCs as described previously (17). The cytokine profiles of T cell clones were determined as described previously (17). Basophils were isolated and purified to homogeneity (>98% purity) following double Percoll density centrifugation and negative selection using a mixture of mAbs (CD2, CD3, CD14, CD16, CD24, CD34, CD36, CD45RA, CD56, glycophorin) and magnetic colloid beads (StemCell Technologies) as described previously (18). Basophils were activated by stimulation with anti-IgE Abs as previously described (18). Human mast cells were generated from umbilical cord blood as described previously (19). Briefly, the mononuclear cell fraction was obtained by centrifugation and suspended in modified DMEM containing 10 μg/ml insulin, 10 μg/ml transferrin, 10 μM 2-ME, 25 mM HEPES, 2.6 ng/ml NaSeO3, 5% FCS, human recombinant stem cell factor at 80 ng/ml (Amgen, Thousand Oaks, CA), human rIL-6 at 50 ng/ml (Amgen), and PGE2 (Sigma, St. Louis, MO) at 300 nM. Cells were used when conformity in morphology and shape of the cells is reached. Purity of mast cells in the preparations was always >95%. Mast cells were activated using myeloma IgE alone (1 μg/ml) or a combination of myeloma IgE (1 μg/ml) and anti-human IgE (1 μg/ml; kindly provided by Dr. R. Hamilton, Johns Hopkins University, Baltimore, MD). Mast cells were sensitized overnight with myeloma IgE at 37°C before challenge with anti-IgE.

ML-1 gene expression was also assayed from bronchoalveolar lavage (BAL)4 cells of four asthmatic patients challenged with either allergen (ragweed, 100 protein nitrogen units) or saline control as described previously (20). The BAL cells were collected 19 h after challenge. Total RNA was isolated from 1) ragweed-activated PBMCs (5 × 106 cells; 6 h after stimulation); 2) peripheral blood CD4+ (1 × 106 cells), CD8+ T cells (5 × 105 cells), and monocytes (1 × 106 cells); all three cell types were stimulated with PMA (100 ng/ml)/ionomycin (2 μg/ml) for 4 h; 3) cloned T cells (2 × 106 cells; 6 h after ragweed allergen stimulation); 4) basophils (2 × 106 cells; 4 h after stimulation); and 5) mast cells (2 × 106 cells; 6 h after stimulation) using RNAzol B according to the manufacturer’s instructions (Tel-Test, Friendswood, TX). cDNAs were synthesized from 500 ng of total RNA in the presence of Moloney murine leukemia virus reverse transcriptase (1 U/reaction; Sigma), oligo(dT) primer, and reaction buffer at 42°C for 90 min, followed by PCR. Each cDNA sample was amplified (30 cycles of 1 min at 95°C, 1 min at 52°C, and 1 min at 72°C) using ML-1 and IL-17 sequence-specific primers (see above). The sequences of primers for a housekeeping gene, G3PDH, were as follows: forward, 5′-ACCACAGTCCATGCCATCAC-3′; reverse, 5′-TCCACCACCCTGTTGCTGTA-3′. The expected size for the ML-1 PCR product is 220 bp, for the IL-17 is 462 bp, and for the G3PDH is 450 bp.

Recombinant ML-1-His fusion proteins

The coding sequence of ML-1 was amplified by PCR and subcloned into the BamHI and XbaI sites of pcDNA 3.1 (Invitrogen, Carlsbad, CA) to generate a C-terminal fusion gene with the His and c-Myc tags. The vector pcDNA 3.1 was transfected into COS-7 cells by an Effectene reagent (Qiagen, Chatsworth, CA) according to the manufacturer’s instructions. Two days after transfection, the supernatants were concentrated over Centricon-10 columns (Amicon, Beverly, MA) and subjected to affinity purification by Ni-NTA agarose beads (Qiagen) for His-tagged proteins. To examine the protein expression, SDS-PAGE analysis was performed on the affinity-purified recombinant proteins under a reducing condition, followed by Western blot analysis using anti-His mAb (Santa Cruz Biotechnology, Santa Cruz, CA).

Analysis of ML-1-induced gene expression

Primary bronchial epithelial (PBE) cells were purchased from Clonetics (San Diego, CA). PBE cells were cultured according to the manufacturer’s instructions. The cells were treated with IL-17 (10 or 100 ng/ml), or ML-1 (10 or 100 ng/ml), or a control His-tagged protein (Positope, 10 or 100 ng/ml; Invitrogen). The affinity-purified control His protein was dissolved in the same buffer as ML-1. Total RNA was extracted using RNeasy (Qiagen) from 1 × 106 cells 4 h after stimulation or exchange of medium. The protocol for cDNA synthesis was the same as described above. For PCR, the sequences of PCR primers were based on the human IL-8 cDNA sequence. The sequences of PCR primers for IL-8 were: forward, 5′-TCTGCAGCTCTGTGTGAAG-3′; reverse, 5′-TAATTTCTGTGTTGGCGCA-3′; IL-6: forward, 5′- ATGAACTCCTTCTCCACAAGCGC-3′, reverse, 5′-GAAGAGCCCTCAGGCTGGACTG-3′. Also, gene expression for two chemokines, eotaxin and RANTES, was also analyzed. The primers for eotaxin and RANTES are as follows: eotaxin: forward, 5′-ACAGGAGAATCACCAGTGGC-3′, reverse, 5′-ACTTCATGGAATCCTGCACC-3′; RANTES: forward, 5′-CGCTGTCATCCTCATTGCTA-3′, reverse, 5′-TGATGTACTCCCGAACCCAT-3′. The amplification reaction was performed for 23 cycles with denaturation at 94°C for 45 s, annealing at 56°C for 45 s, and extension at 72°C for 45 s. PCR products were detected by ethidium bromide staining and normalized by the intensity of an amplified housekeeping gene, G3PDH (see above). The expected size for IL-8 was 154 bp and for IL-6, 628 bp. IL-6 and IL-8 protein levels in the collected supernatants were determined with a commercially available ELISA kit (BioSource International, Camarillo, CA) according to the manufacturer’s instruction.

Analysis of ICAM-1 surface expression by flow cytometry

PBE cells were treated with IL-17 (100 ng/ml), ML-1 (100 ng/ml), or a control His protein (Positope, 100 ng/ml) for 48 h. The cells were harvested following treatment with 0.025% trypsin-0.02% EDTA at 37°C for 6 min and suspended in PBS containing 2% FBS and 0.02% sodium azide. Briefly, 106 cells were incubated with a mouse anti-human ICAM-1 mAb (R&D Systems, Minneapolis, MN) on ice for 30 min. After three washes with PBS, the cells were incubated with FITC-conjugated goat anti-mouse IgG (Bio-Rad, Hercules, CA) on ice for 30 min. After three additional washes with PBS, the cells were resuspended in PBS and immediately analyzed with a FACScan flow cytometer (BD Biosciences, Mountain View, CA). In control samples, staining was performed using isotype-matched control Abs. The mean fluorescence intensity (MFI) was expressed as mean ± SD (n = 5) by subtracting the mean background fluorescence.

Effect of IL-17 on ML-1-induced ICAM-1 gene expression

To assess whether ML-1 interacts with IL-17R, competitive inhibition experiments were performed using the level of ICAM-1 gene expression as a functional readout, since IL-17 by itself is unable to induce ICAM-1 expression. PBE cells were treated with either medium, ML-1 (100 ng/ml) alone, or in combination with varying doses (10–2000 ng/ml) of IL-17 (R&D Systems for 4 h, and total RNAs were extracted from each sample as described above. RT-PCR was performed using the primers based on the human ICAM-1 cDNA sequence. The sequences of primers for ICAM-1 were: forward, 5′-CAGAGGTTGAACCCCACAG-3′, reverse, 5′-CCTCTGGCTTCGTCAGAAT-3′. The amplification was performed for 25 cycles with denaturation at 94°C for 45 s, annealing at 55°C for 45 s, and extension at 72°C for 45 s. PCR products were detected by ethidium bromide staining, and normalized by the intensity of an amplified housekeeping gene, G3PDH (see above). The expected size for ICAM-1 was 196 bp.

Data analysis

All data are expressed as mean ± SD. Differences between the groups were analyzed by a one-way ANOVA analysis with the Scheffe F test by using the StatView program (SAS Institute, Cary, NC) on an Apple computer (Apple Computer, Cupertino, CA). Differences were considered to be significant at p < 0.05.


Full-length cDNA sequence of the ML-1 gene

As part of a positional cloning study of a region on chromosome 6p for susceptibility gene discovery for an inherited disease, a potential coding region sequence with homology to human IL-17 was identified from a genomic DNA clone, PAC108C2* (Sanger Center Database, http://www.sanger.ac.uk), using a GenScan prediction program. The predicted expressed sequence with a centromeric-telomeric orientation was composed of 2 exons of 221 and 238 bp, respectively. RT-PCR and sequencing analysis of activated, human allergen-specific T cell clones confirmed the predicted sequence and the splicing sites between the exons. However, the open reading frame utilizes a start codon 129 bp 3′ to the predicted start site, encompassing a 92-bp segment in the first exon. A full-length cDNA 947 bp in length was obtained using both 5′- and 3′-RACE, revealing a transcription start site 346 bp upstream of the start codon and a poly(A) sequence 271 bp 3′ to the stop codon.

Homology searches using the BLAST program shows a significant degree of homology between the second exon of the ML-1 gene and the third exon of IL-17. Alignment of the predicted amino acid sequence of ML-1 with sequences of IL-17 and the other members of the IL-17 family shows that while there is the highest overall amino acid sequence homology (70%) between ML-1 and IL-17, there is only 20% amino acid identity between ML-1 and the three other family members (Fig. 1⇓). The alignment shows several conserved amino acids including a tryptophan residue and four cysteines in the C-terminal half of the proteins.


Amino acid sequence alignment of ML-1, IL-17, IL-17C, and IL-17B. The residues identical to the ML-1 sequence are colored, and conserved cysteine residues are indicated by arrows. The GenBank accession number for the ML-1 cDNA sequence is AF332389.

Expression of the ML-1 gene

The expression of ML-1 in various cell types and human tissues was examined using PCR and the Rapid-Scan gene expression panels for human tissues. Table I⇓ shows the tissue expression of ML-1 and IL-17. ML-1 was strongly expressed in liver, lung, spleen, placenta, adrenal gland, ovary, and fetal liver. Interestingly, ML-1 expression in liver, lung, ovary, and fetal liver was unique when compared with IL-17 (Table I⇓). In addition, although the expression of ML-1 was not detected in unstimulated cells, with the exception of mast cells (Fig. 2⇓A), increased ML-1 expression was clearly evident in six different cell types after activation (Fig. 2⇓B). Those cell types included ragweed allergen-stimulated PBMCs, ragweed allergen-specific T cell clones with different cytokine phenotypes (Th0 (clone 12), Th1 (clone 2B7), and Th2 (clone 2D2)), activated basophils, and activated mast cells (Fig. 2⇓B). Also, expression of ML-1 was found in activated peripheral blood CD4+ T cells, but not CD8+ and monocytes (Fig. 2⇓C). In contrast, IL-17 gene expression was found in only activated peripheral blood CD4+ T cells, Th0 clones, and PBMCs (Fig. 2⇓).


Expression patterns of the ML-1 gene. RT-PCR analysis of ML-1 and IL-17 expression in unstimulated (A) and activated (B) cell types as indicated. Expression of ML-1 and IL-17 genes in resting or activated peripheral blood CD4+, CD8+ T cells, and monocytes (C). Expression of ML-1 and IL-17 genes at sites of airway inflammation (D). Detection of gene expression in BAL cells from four asthmatic patients (BAL #638, #646, #688, and #1081) challenged with allergen (Ag) and saline control (NS). Amplification of G3PDH was performed as a positive control.

This unique expression pattern of the ML-1 gene in activated Th2 cells, a cell type known to be associated with allergic asthma, prompted us to determine the in vivo relevance of ML-1 gene expression. We performed analyses of gene expression in BAL cells from asthmatic subjects challenged with allergen or saline control. Fig. 2⇑C shows representative data demonstrating that while no detectable expression of ML-1 was seen in the BALs from saline-challenged sites, ML-1 gene expression was clearly demonstrated in the BALs from allergen-challenged sites of all four study subjects. In contrast, no IL-17 transcripts were detected in the BALs of subjects challenged with allergen (Fig. 2⇑D).

Table I.

Tissue expression of ML-1 and IL-17a

Induction of IL-6, IL-8, and ICAM-1 expression in PBE cells

To assess the biological functions of ML-1, His-tagged ML-1 protein was expressed in COS-7 cells and affinity purified (Fig. 3⇓A). PBE cells were treated with either affinity-purified ML-1 or a His-tagged control protein and assayed for IL-6, IL-8, and ICAM-1 expression. Similar to IL-17, ML-1 at two different doses, 10 and 100 ng/ml, enhanced IL-6 and IL-8 transcripts (Fig. 3⇓, B and C) and protein expression for 48 h in PBE cells (Fig. 3⇓, D and E; for IL-8, 1724.2 ± 132.9 and 4371.4 ± 280.5 pg/ml, respectively; for IL-6, 599.6 ± 19.1 and 1005.3 ± 55.6 pg/ml, respectively). No increase of IL-6 and IL-8 expression was seen in cells treated with either His-tag control protein or medium control. Furthermore, ML-1 did not induce the gene expression for two CC chemokines, eotaxin and RANTES (Fig. 3⇓F).


Expression of ML-1 protein in COS-7 cells (A). The supernatants from COS-7 cells transfected with a gene construct containing the full-length ML-1 sequence and a tag sequence encoding poly(His) peptide. ML-1-His fusion proteins were purified by His affinity column, run on 8–16% SDS-PAGE under reducing conditions, and analyzed by Western blot using anti-His mAb. Lane 1, A mock-transfected sample, and lane 2, purified protein from an ML-1-transfected sample. The location of a protein size marker, 19.5 kDa, is indicated. Effect of ML-1 and IL-17 on IL-8 (B) and IL-6 (C) gene expression. PBE cells were treated with medium alone, rIL-17 (100 ng/ml), purified His-tagged protein (Positope, 100 ng/ml), or ML-1 (100 ng/ml) as indicated. The cells were harvested 4 h after stimulation, and total RNA was extracted and subjected to RT-PCR using IL-8, IL-6, and G3PDH primers. The relative level of IL-8 expression is normalized by the level of G3PDH. Effect of ML-1 and IL-17 on IL-8 (D) and IL-6 (E) protein production. PBE cells were treated as indicated, and the supernatants were harvested 48 h after stimulation. The experiment was conducted five times. ∗, p < 0.05. Analysis of gene expression for eotaxin and RANTES (RAN) in PBE cells in the presence (+) or absence (−) of 100 ng/ml ML-1 (F).

It has been previously shown that IL-17 alone is not able to induce ICAM-1 expression on airway epithelial cells (12), but synergizes with IFN-γ for the induction of ICAM-1 expression on epithelial cells. Up-regulation of ICAM-1 expression is associated with airway inflammation seen in bronchial asthma (13). To examine whether ML-1 is able to induce ICAM-1 expression on PBE cells, surface expression of ICAM-1 on PBE cells was measured and the effects of ML-1 vs IL-17 were compared. Fig. 4⇓A demonstrates that ICAM-1 was constitutively, but weakly, expressed on PBE cells (MFI = 12.26 ± 1.77) and IL-17 (100 ng/ml) alone did not affect ICAM-1 surface expression at 48 h (MFI = 11.07 ± 1.22). The supernatants of COS-7 cells alone or in combination with IL-17 did not induce ICAM-1 expression (data not shown). In contrast, ML-1 (100 ng/ml) significantly induced ICAM-1 expression at 48 h (MFI = 31.42 ± 4.39) (p < 0.05) when compared with those seen in cells stimulated with IL-17 and a His-tagged protein (MFI = 12.99 ± 2.09). Furthermore, similar patterns of differential effects on ICAM-1 gene expression were also observed (data not shown). These findings suggest a distinct function of ML-1, despite the fact that IL-17 and ML-1 share a significant degree of sequence homology. Furthermore, the functional disparity between IL-17 and ML-1 in the induction of ICAM-1 expression suggests differential interaction with cellular receptors. To assess whether ML-1 interacts with IL-17R, competitive inhibition experiments were performed by measuring the level of ML-1-induced ICAM-1 gene expression in the presence of varying concentrations of IL-17. As shown in Fig. 4⇓B, ML-1 at 100 ng/ml markedly induced ICAM-1 expression, and the level of gene expression normalized by the level of G3PDH expression was not inhibited by the inclusion of IL-17, even at a 20-fold higher dose (2000 ng/ml). These differential biological effects between ML-1 and IL-17 suggest that these two cytokines may signal via different cell surface receptors. Further work is needed to identify the nature of the putative ML-1R.


Effect of ML-1 on ICAM-1 expression on PBE cells. A, Flow cytometry analysis. PBE cells were treated for 48 h under different conditions: medium only, cells treated with rIL-17 (100 ng/ml), cells treated with purified His-tagged protein (Positope, 100 ng/ml), or cells treated with ML-1 (100 ng/ml) as indicated, and the MFI of ICAM-1 surface expression was measured. The experiment was conducted five times. ∗, p < 0.05. B, ML-1-induced ICAM-1 gene expression. PBE cells were treated with either medium, ML-1 (100 ng/ml) alone, or in combination with different doses of IL-17 as indicated. The cells were harvested 4 h after stimulation, and total RNA was extracted and subjected to RT-PCR. The relative level of ICAM-1 expression is normalized by the level of G3PDH. The experiment was conducted three times, and the results shown are from a representative experiment. ∗, p < 0.05.


We report herein the identification and characterization of a new cytokine with a distinct expression pattern and function when compared with other members of the IL-17 gene family. Although members of the IL-17 gene family are classified based on amino acid sequence similarity, the genes are selectively expressed in different tissues and are dispersed in the genome (1, 14, 15). Although IL-17 expression is restricted to activated PBMCs and activated Th0 cells, but not in activated Th2 cells (Fig. 2⇑B; Refs. 1, 2, 3, 21), ML-1 is expressed in various cell types and tissues. Its distribution is much wider than that of IL-17, suggesting ML-1 has more biological functions. In contrast to ML-1 and IL-17, two other members of the IL-17 family, IL-17B and IL-17C, were not detected in activated CD4+ T cells (14, 15). Moreover, among the IL-17 family members, ML-1 shows the highest homology with human IL-17. It is noted, however, that contrary to a previous report, the IL-17 gene is localized on the same genomic DNA clone (GenBank accession no. AL391221) ∼50 kb telemeric to ML-1 sequence and both genes are in a tail-to-tail orientation (data not shown), suggesting a potential gene duplication event.

Two other members of the IL-17 gene family, IL-17B and IL-17C, each share ∼27% amino acid identity with IL-17 (14, 15). IL-17B mRNA is expressed in adult pancreas, small intestine, and stomach, whereas IL-17C mRNA is not detected in the same set of adult tissues. Interestingly, no expression of IL-17B or IL-17C mRNA is found in activated T cells (15). Both IL-17B and IL-17C stimulate the release of TNF and IL-1 from a monocytic cell line, THP-1. However, IL-17B and IL-17C are not able to stimulate IL-6 production from human fibroblasts and do not bind to the human IL-17R. Furthermore, a newly discovered member of the IL-17 gene family, IL-17E, has been shown to induce activation of NF-κB and IL-8 via a distinct receptor (16). Taken together, these results and other published studies suggest the existence of a family of IL-17-related cytokines differing in their patterns of tissue/cell expression and in their potential roles in immunologic responses.

ML-1 and IL-17 showed the same potency in the induction of IL-6 and IL-8, suggesting that ML-1 is involved in neutrophil recruitment into the airway such as IL-17 (11). The level of IL-8 is increased in inflammatory airway diseases, such as bronchial asthma, chronic obstructive pulmonary disease, and cystic fibrosis (22, 23, 24). There is a strong correlation between the level of IL-8 and the increase in neutrophil numbers (22). An anti-IL-8 mAb inhibited neutrophil chemotaxis in patients with chronic obstructive pulmonary disease or cystic fibrosis (24). In addition, bronchial epithelial cells are an important cell source of IL-8 and may thereby control neutrophil influx in the airway (25). Therefore, IL-8 release in bronchial epithelial cells may play a crucial role in modulating neutrophil-associated airway inflammation.

Although IL-17 and ML-1 induce IL-8 expression from PBE cells, IL-17 fails to induce ICAM-1 expression in human bronchial epithelial cells (Fig. 4⇑A and Ref. 12). By comparison, ML-1 markedly induced ICAM-1 expression. It has been shown that ICAM-1 expression is increased in airway diseases, such as bronchial asthma (13). In particular, a high level of ICAM-1 expression, along with inflammatory cell infiltration, has been demonstrated in bronchial biopsies from both stable asthmatics and subjects after allergen challenge (13, 26). Moreover, allergen challenge up-regulated ICAM-1 expression in airway epithelium, correlating with eosinophil infiltration. Increased expression of ML-1, but not IL-17, is also seen in patients with asthma following allergen challenge. Moreover, expression of ML-1 is detected in activated Th2 cells, basophils, and mast cells, three important cell types involved in allergic responses. These findings suggest, therefore, that ML-1 may be involved in the expression of airway inflammation by, at least in part, facilitating leukocyte recruitment and activation via the induction of IL-8 and ICAM-1 in airway epithelium. Further investigation is necessary to clarify the mechanism(s) by which ML-1 is involved in normal physiology, asthma, and other inflammatory diseases.


We thank Beverly Plunkett and Eva Ehrlich for their excellent technical assistance.


  • ↵1 This work was supported by National Institutes of Health Grants AI-20059 and AI-34002 (to S.K.H.).

  • ↵2 M.K. and L.F.O. contributed equally to this work.

  • ↵3 Address correspondence and reprint requests to Dr. Shau-Ku Huang, Johns Hopkins Asthma and Allergy Center, 5501 Hopkins Bayview Circle, Baltimore, MD 21224-6801. E-mail address: skhuang{at}mail.jhmi.edu

  • ↵4 Abbreviations used in this paper: BAL, bronchoalveolar lavage; PBE, primary bronchial epithelial; MFI, mean fluorescence intensity.

  • Received May 14, 2001.
  • Accepted August 9, 2001.
  • Copyright © 2001 by The American Association of Immunologists


Our History

In the early 70’s a group of former members of New York Holy Cross Church living in Westchester came together for the planning of a new parish in Westchester. Menas Daderian presented the wish of the group to then Primate Bishop Torkom Manoogian, who welcomed the wish, writing on June 8, 1973: “In response to your letter of May 18, and the highly commendable sentiments for the establishment of a new parish in Westchester, NY, we are pleased to appoint a group of eighteen persons to serve as a temporary parish council… Consideration will be given to the assignment of a permanent priest when the parish is able to support a pastor. Until such time we will tentatively assign Fr. Arshen Aivazian as the spiritual advisor to the Parish Council. Perhaps the most important matter in this stage of your organization is the raising of funds.” The Primate appointed the following individuals as the Executive Committee of the temporary Parish Council: Menas Daderian, Chairman; George Davidian, Vice Chairman; Elma Davidian, Corresponding Secretary; Agnes Babian, Recording Secretary; George Sanossian, Treasurer; David Davidian, Publicity.

In the meantime, three seminarians of st. Nersess Seminary, together with their Dean, Deacon Hovhannes Kasparian and his wife Dawn, met in the home of Edward and Barbara Dorian to compile a mailing list of Armenians from the Westchester telephone book. Soon some parishioners and St. Nersess seminarians joined to teach Armenian language and Sunday school classes using the facilities of St. Vladimir Seminary in Crestwood. Monthly gatherings of religious and cultural programs for adults were also initiated.

In the spring of 1974, the Primate appointed Deacon Hovhannes as spiritual advisor to the new parish. The first Parish Assembly was held and the first Parish Council was elected on March 22, 1974. On May 19, 1974 Deacon Hovannes was ordained into the priesthood as Father Karekin and assumed part-time pastorate of the new parish while maintaining part-time responsibility at St. Nersess. He became fulltime parish priest in 1978, when Edward S. Dorian Sr. was the Chairman of the parish council.

Thereafter, many facets of church life began to flourish. A choir was established and various church organizations were formed: Mr. and Mrs. Group, Youth Group, Senior & Junior ACYOA, Women’s Guild, Men’s Club, Senior (Avaks) and later the Hye Steppers. All the groups began to have various interesting social and educational programs which attracted many new members.

Since the first Divine Liturgy, celebrated by Fr. Carnig Hallajian at the Manhattanville College Chapel on October 25, 1973, the growing number of congregants have participated in worship in seven different locations, including the Seventh Day Adventist Church in Yonkers and St. Andrew’s Episcopal Church in Hartsdale.

In 1980, the congregation purchased a seven-acre parcel of land with a large private house in Purchas, NY. Under the steadfast leadership of George Davidian, the legal advice of Judge Vincent Gurahian and the strong support of Suren and Virginia Fesjian, a building on the property was transformed into a church and named St. James (Soorp Hagop) with Mr. Fesjian serving as Gnkahayr. In 1985 the parish decided to initiate a search for a site to build a new parish with the initiative of Council Chairman Dr. Jack Hachigian. A fundraising campaign was launched with a dinner-dance, when two brothers Simon and Cicek and Hayik and Vartouhi Tutak made a donation of $300,000 for the construction of the new fellowship hall. The Parish has always been grateful to the Fesjians and the Tutak brothers for their generous donations at crucial stages in the life of this community.

Shepherded by the Pastor, the parish grew significantly during its 17 years in Purchase, attracting active parishioners from all parts of greater Westchester County and Southern Connecticut. From the early years, when our first choir director was Deacon Vazken Asadourian (now Archbishop Avak, Primate of the Diocese of IRAQ) to his successor Alfred Yeznaian our choir began to grow. Worship services were enhanced with the appointment of Raffi P. Sevadjian as our choir director and the ordination of John Vakassian and Mark Kapikian as senior deacons. St. James became a dynamic parish responding to the spiritual, cultural, social and communal needs of hundreds of Armenians in the area.

The hard work of many members enabled the parish to meet its financial obligations. Among the others, the service of Garo Bashian with the help of his wife Annette was instrumental in raising funds by initiating and conducting, for many years, the successful Oriental Rug sales. The annual picnics have become not only occasions for enjoyable fellowship for hundreds of parishioners from our own and sister parishes, but also another dependable source of income to meet the annual budget. In recent years, aided by a cadre of dedicated parishioners, Michael Guroian has been the tireless chairman of many successful parish picnics.

Several other ventures and undertakings have helped the parish remain financially solvent; members have generously given of their time, talent and treasure for the nurturing of the church. The volunteers of St. James, from the beginning, have devoted their unique talents to bring our initial idea of a parish to fruition, their labors have included cleaning, painting, plumbing, repairing, installing play equipment, mowing grass, teaching children, singing in the choir, making signs for special events, donating homemade baked goods, installing air conditioning and running dances, bazaars, raffles and tag sales, selling lehmejune and luleh kabob, moving furniture, decorating for holidays, providing legal advice, collectin funds for earthquake victims and hundreds of other selfless deeds all for the good of the parish. Throughout is history of over one quarter of a century, the parish has/had many educational, cultural and social programs. Valuable service has been rendered to the Armenian educational growth of the children of the parish through the Armenian School where dedicated teachers have been serving close to two decades. The Sunday School also grew from a handful of students and has been functioning smoothly throughout the years thanks to the dedication of many teachers.

The growth of the community and its expanding needs necessitated the purchase of new property. The present site in White Plains was purchased in 1993 after a relentless search by a team headed by Edward Essayan. Our Primate, the Most Rev. Archbishop Khajag Barsamian blessed the property on September 16, 1995. On January 14, 1996 the parish was privileged to have His Holiness Karekin I, Catholicos of All Armenians, bless the ten blocks of “TOOF” stone from Armenia as the cornerstone of the new church.

In an unprecedented act of Christian giving and piety, Mr. and Mrs. Hayik Tutak made a generous gift of $1,000,000.00 in order to move the construction of our new magnificent church complex forward. This princely gesture further encouraged parishioners to participate in the fundraising campaign, formerly chaired by Ara Momjian, including a donation of $100,000.00 by the late Anahid Birnbaum.

In April 1996, the parish sold the property in Purchase and held a farewell banquet at which time the Primate honored Mr. Fesjian as the founding Grand Benefactor of Soorp Hagop Church and welcomed Mr. and Mrs. Hayik Tutak as the new Grand Benefactors. For over nineteen months, during the construction of the church complex, worship services and Sunday School classes were held at St. Nersess Seminary while the parish house on the new property were utilized for offices, Armenian School classes and other church group activities.

A Timeline of Our Parish and Church

1971: A small group of Armenians living in Westchester move on the concept of forming an Armenian Church of Westchester. Dn. Hovhannes Kasparian, Dean of St. Nersess Seminary (at St. Vladimir’s Seminary) appointed spiritual advisor to the Armenians in Westchester and invites community to monthly cultural/educational gatherings at St. Vladimir’s Seminary, where Armenian language and religious classes are initiated. Divine Liturgy celebrated at St. Vladimir’s by Fr. Arshen Aivazian.

1973: Westchester Armenians receive permission to form a new parish from Abp. Manoogian of the Armenian Diocese, who appoints a Temporary Parish Council, with Menas Daderian as its Chair. This Council sends a letter to all Armenians living in the area soliciting interest in establishing and Armenian Church in Westchester. First Divine Liturgy in Westchester is celebrated at Manhattanville College Chapel in October.

1974: Dn. Hovhannes is ordained as Fr. Karekin and becomes part-time pastor of the new parish. The first elected Parish Council takes office, with Edward S. Dorian as Chair, who also takes on the roles of Fundraising Chair, and first elected Diocesan Delegate. During these formative years, parishioners worship in 7 different locations.

1975: The Parish Council forms a Building Fund Committee; Women’s Guild is organized with 15 members. The church newsletter is named Hope.

1977: Suren D. Fesjian matches the donations of the Bi-centennial Building Drive. Menas Daderian is appointed chair fo the Site Finding Committee.

1978: Fr. Kasparian assumes full pastorate of the church. Stewardship program is established.

1979: Purchase of Harrison property for $275,000.00 subsequently approved for use as a church. Volunteer parishioners, headed by George Davidian, convert 3-car garage to a temporary chapel. George and Elma Davidian honored as Couple of the Year, starting an annual tradition now known as St. Gregory Award Program.

1980: First Badarak held at the new church property on June 8. The church is named St. James/Soorp Hagop, with Suren D. Fesjian, the major donor, as Godfather.

1986: The church begins a building initiative. A generous pledge by brothers Simon and Hayik Tutak, as sponsors of a new fellowship hall, boosts Building Fund Campaign.

1990: 10th Anniversary of St. James/Soorp Hagop church.

1991: Parish Council Chairman Jack Hachigian spearheads a movement to expand to a larger facility.

1993: New site is purchased for the permanent church in White Plains, NY. Building phase continues over the next 5 years.

1997: Mr. and Mrs. Hayik Tutak gift one million dollars toward building of the church complex, allowing the project to reach completion.

1998: Consecration of St. Gregory the Enlightener Church by Abp. Khajag Barsamian, presided over by His Holiness Karekin I, Catholicos of All Armenians.

1999: Parish celebrates 1st Anniversary of new church and 25th Anniversary of pastor’s ordination. The new Sunday School and Armenian School facilities are dedicated.

2000: The Manoogian Library opens, sponsored by Louise Manoogian Simone. New playground built with donation by Raffi and Felicia Atamian. Der Karekin assigned by Abp. Barsamian to head Diocesan Pastoral Support Ministry and Pastoral Internship Program at St. Gregory.

2001: First Ecumenical Service at St. Gregory. 30th Anniversary of our Armenian parish in Westchester; 3rd Anniversary of St. Gregory the Enlightener Church; 1700th Anniversary of the Christianization of Armenia. Sister church St. Gregory at Khor Virab (Armenia) adopted – black earth from Khor Virab and a candlestick permanently placed on center altar as relics from the oldest place associated with the life of the Enlightener.

2002: White House is renovated.

2003: Expansion of Sunday School/Armenian School classrooms. Church mortgage satisfied, and burned. White House Thrift Shoppe, co-chaired by Alice Basmajian and Mary Sohigian, established by the Women’s Guild. St. Gregory’s Men’s Club revitalized under the leadership of Harry Keleshian.

2004: 102nd Diocesan Assembly hosted at St. Gregory.

2006: First pilgrimage to Armenia instituted by Armenian School graduates.

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