Antagonism of Ang-Tie2 and Dll4-Notch signaling has opposing effects on tumor endothelial cell proliferation, evidenced by a new flow cytometry method.

Sustained angiogenesis is essential for tumor growth as it provides the tumor with a network of blood vessels that supply both oxygen and essential nutrients. Limiting tumor-associated angiogenesis is a proven strategy for the treatment of human cancer. To date, the rapid detection and quantitation of tumor-associated endothelial cell (TAEC) proliferation has been challenging, largely due to the low frequency of endothelial cells (ECs) within the tumor microenvironment. In this report, we address this problem using a new multiparametric flow cytometry method capable of rapid and precise quantitation of proliferation by measuring bromodeoxyuridine (BrdUrd) uptake in mouse TAECs from established human tumor xenografts. We determined the basal proliferation labeling index of TAECs in two human tumor xenografts representing two distinct histologies, COLO 205 (colorectal cancer) and U-87 (glioblastoma). We then investigated the effects of two large-molecule antiangiogenic agents targeting different biochemical pathways. Blocking angiopoietin-Tie2 signaling with the peptide-Fc fusion protein, trebananib (AMG 386), inhibited proliferation of TAECs, whereas blocking Dll4-Notch signaling with an anti-Dll4-specific antibody induced hyperproliferation of TAECs. These pharmacodynamic studies highlight the sensitivity and utility of this flow cytometry-based method and demonstrate the value of this assay to rapidly assess the in vivo proliferative effects of angiogenesis-targeted agents on both the tumor and the associated vasculature.

Epo receptors are not detectable in primary human tumor tissue samples.

Erythropoietin (Epo) is a cytokine that binds and activates an Epo receptor (EpoR) expressed on the surface of erythroid progenitor cells to promote erythropoiesis. While early studies suggested EpoR transcripts were expressed exclusively in the erythroid compartment, low-level EpoR transcripts were detected in nonhematopoietic tissues and tumor cell lines using sensitive RT-PCR methods. However due to the widespread use of nonspecific anti-EpoR antibodies there are conflicting data on EpoR protein expression. In tumor cell lines and normal human tissues examined with a specific and sensitive monoclonal antibody to human EpoR (A82), little/no EpoR protein was detected and it was not functional. In contrast, EpoR protein was reportedly detectable in a breast tumor cell line (MCF-7) and breast cancer tissues with an anti-EpoR polyclonal antibody (M-20), and functional responses to rHuEpo were reported with MCF-7 cells. In another study, a functional response was reported with the lung tumor cell line (NCI-H838) at physiological levels of rHuEpo. However, the specificity of M-20 is in question and the absence of appropriate negative controls raise questions about possible false-positive effects. Here we show that with A82, no EpoR protein was detectable in normal human and matching cancer tissues from breast, lung, colon, ovary and skin with little/no EpoR in MCF-7 and most other breast and lung tumor cell lines. We show further that M-20 provides false positive staining with tissues and it binds to a non-EpoR protein that migrates at the same size as EpoR with MCF-7 lysates. EpoR protein was detectable with NCI-H838 cells, but no rHuEpo-induced phosphorylation of AKT, STAT3, pS6RP or STAT5 was observed suggesting the EpoR was not functional. Taken together these results raise questions about the hypothesis that most tumors express high levels of functional EpoR protein.

Novel anti-c-Mpl monoclonal antibodies identified multiple differentially glycosylated human c-Mpl proteins in megakaryocytic cells but not in human solid tumors.

Thrombopoietin and its cognate receptor, c-Mpl, are the primary molecular regulators of megakaryocytopoiesis and platelet production. To date the pattern of c-Mpl expression in human solid tumors and the distribution and biochemical properties of c-Mpl proteins in hematopoietic tissues are largely unknown. We have recently developed highly specific mouse monoclonal antibodies (MAb) against human c-Mpl. In this study we used these antibodies to demonstrate the presence of full-length and truncated human c-Mpl proteins in various megakaryocytic cell types, and their absence in over 100 solid tumor cell lines and in the 12 most common primary human tumor types. Quantitative assays showed a cell context-dependent distribution of full-length and truncated c-Mpl proteins. All forms of human c-Mpl protein were found to be modified with extensive N-linked glycosylation but different degrees of sialylation and O-linked glycosylation. Of note, different variants of full-length c-Mpl protein exhibiting differential glycosylation were expressed in erythromegakaryocytic leukemic cell lines and in platelets from healthy human donors. This work provides a comprehensive analysis of human c-Mpl mRNA and protein expression on normal and malignant hematopoietic and non-hematopoietic cells and demonstrates the multiple applications of several novel anti-c-Mpl antibodies.

Cinacalcet attenuates hypercalcemia observed in mice bearing either Rice H-500 Leydig cell or C26-DCT colon tumors.

Excessive secretion of parathyroid hormone-related protein (PTHrP) by tumors stimulates bone resorption and increases renal tubular reabsorption of calcium, resulting in hypercalcemia of malignancy. We investigated the ability of cinacalcet, an allosteric modulator of the calcium-sensing receptor, to attenuate hypercalcemia by assessing its effects on blood ionized calcium, serum PTHrP, and calcium-sensing receptor mRNA in mice bearing either Rice H-500 Leydig cell or C26-DCT colon tumors. Cinacalcet effectively decreased hypercalcemia in a dose- and enantiomer-dependent manner; furthermore, cinacalcet normalized phosphorus levels, but did not affect serum PTHrP. Ribonuclease protection assay results demonstrated presence of PTHrP receptor, but not calcium-sensing receptor mRNA in C26-DCT tumors. The mechanism by which cinacalcet lowered serum calcium was investigated in parathyroidectomized rats (i.e., without PTH) made hypercalcemic by PTHrP. Cinacalcet attenuated PTHrP-mediated elevations in blood ionized calcium, which were accompanied by increased plasma calcitonin. Taken together these results suggest that the cinacalcet-mediated decrease in serum calcium is not the result of a direct effect on tumor cells, but rather is the result of increased calcitonin release. In summary, cinacalcet effectively reduced tumor-mediated hypercalcemia and corrected hypophosphatemia in mice. Further investigation of cinacalcet for treatment of hypercalcemia of malignancy is warranted.

A protective role for keratinocyte growth factor in a murine model of chemotherapy and radiotherapy-induced mucositis.

PURPOSE: To evaluate the activity of palifermin (rHuKGF) in a murine model of mucosal damage induced by a radiotherapy/chemotherapy (RT/CT) regimen mimicking treatment protocols used in head-and-neck cancer patients. METHODS AND MATERIALS: A model of mucosal damage induced by RT/CT was established by injecting female BDF1 mice with cisplatin (10 mg/kg) on Day 1; 5-fluorouracil (40 mg/kg/day) on Days 1-4, and irradiation (5 Gy/day) to the head and neck on Days 1-5. Palifermin was administered subcutaneously on Days -2 to 0 (5 mg/kg/day) and on Day 5 (5 mg/kg). Evaluations included body weight, organ weight, keratinocyte growth factor receptor expression, epithelial thickness, and cellular proliferation. RESULTS: Initiation of the radiochemotherapeutic regimen resulted in a reduction in body weight in control animals. Palifermin administration suppressed weight loss and resulted in increased organ weight (salivary glands and small intestine), epithelial thickness (esophagus and tongue), and cellular proliferation (tongue and salivary glands). CONCLUSIONS: Administration of palifermin before RT/CT promotes cell proliferation and increases in epithelial thickness in the oral mucosa, salivary glands, and digestive tract. Palifermin administration before and after RT/CT mitigates weight loss and a trophic effect on the intestinal mucosa and salivary glands, suggesting that palifermin use should be investigated further in the RT/CT settings, in which intestinal mucositis and salivary gland dysfunction are predominant side effects of cytotoxic therapy.

CSMD1 is a novel multiple domain complement-regulatory protein highly expressed in the central nervous system and epithelial tissues.

In this study, we describe the identification and in vitro functional activity of a novel multiple domain complement regulatory protein discovered based on its homology to short consensus repeat (SCR)-containing proteins of the regulators of complement activation (RCA) gene family. The rat cDNA encodes a predicted 388-kDa protein consisting of 14 N-terminal CUB domains that are separated from each other by a SCR followed by 15 tandem SCR domains, a transmembrane domain, and a short cytoplasmic tail. This protein is the homolog of the human protein of unknown function called the CUB and sushi multiple domains 1 (CSMD1) protein. A cloning strategy that incorporates the two C-terminal CUB-SCR domains and 12 of the tandem SCR repeats was used to produce a soluble rat CSMD1 protein. This protein blocked classical complement pathway activation in a comparable fashion with rat Crry but did not block alternative pathway activation. Analysis of CSMD1 mRNA expression by in situ hybridization and immunolabeling of neurons indicates that the primary sites of synthesis are the developing CNS and epithelial tissues. Of particular significance is the enrichment of CSMD1 in the nerve growth cone, the amoeboid-leading edge of the growing neuron. These results suggest that CSMD1 may be an important regulator of complement activation and inflammation in the developing CNS, and that it may also play a role in the context of growth cone function.

Suppression of angiogenesis and tumor growth by selective inhibition of angiopoietin-2.

Angiopoietin-2 (Ang2) exhibits broad expression in the remodeling vasculature of human tumors but very limited expression in normal tissues, making it an attractive candidate target for antiangiogenic cancer therapy. To investigate the functional consequences of blocking Ang2 activity, we generated antibodies and peptide-Fc fusion proteins that potently and selectively neutralize the interaction between Ang2 and its receptor, Tie2. Systemic treatment of tumor-bearing mice with these Ang2-blocking agents resulted in tumor stasis, followed by elimination of all measurable tumor in a subset of animals. These effects were accompanied by reduced endothelial cell proliferation, consistent with an antiangiogenic therapeutic mechanism. Anti-Ang2 therapy also prevented VEGF-stimulated neovascularization in a rat corneal model of angiogenesis. These results imply that specific Ang2 inhibition may represent an effective antiangiogenic strategy for treating patients with solid tumors.

The candidate neuroprotective agent artemin induces autonomic neural dysplasia without preventing peripheral nerve dysfunction.

Artemin (ART) signals through the GFR alpha-3/RET receptor complex to support sympathetic neuron development. Here we show that ART also influences autonomic elements in adrenal medulla and enteric and pelvic ganglia. Transgenic mice over-expressing Art throughout development exhibited systemic autonomic neural lesions including fusion of adrenal medullae with adjacent paraganglia, adrenal medullary dysplasia, and marked enlargement of sympathetic (superior cervical and sympathetic chain ganglia) and parasympathetic (enteric, pelvic) ganglia. Changes began by gestational day 12.5 and formed progressively larger masses during adulthood. Art supplementation in wild type adult mice by administering recombinant protein or an Art-bearing retroviral vector resulted in hyperplasia or neuronal metaplasia at the adrenal corticomedullary junction. Expression data revealed that Gfr alpha-3 is expressed during development in the adrenal medulla, sensory and autonomic ganglia and their projections, while Art is found in contiguous mesenchymal domains (especially skeleton) and in certain nerves. Intrathecal Art therapy did not reduce hypalgesia in rats following nerve ligation. These data (1) confirm that ART acts as a differentiation factor for autonomic (chiefly sympathoadrenal but also parasympathetic) neurons, (2) suggest a role for ART overexpression in the genesis of pheochromocytomas and paragangliomas, and (3) indicate that ART is not a suitable therapy for peripheral neuropathy.

Effect of recombinant human keratinocyte growth factor (rHuKGF) on the immunopathogenesis of intestinal graft-vs.-host disease induced without a preconditioning regimen.

We studied the effect of rHuKGF on acute, lethal graft- vs.-host disease (GVHD) in the C57BL/6-->(C57BL/6 X DBA/2)F(1)-hybrid model. rHuKGF-treated recipients did not develop intestinal GVHD despite elevated levels of intestinal NO and TNF alpha, did not develop endotoxemia, and did not die. LPS augmented serum TNF alpha release and intestinal NO production, but did not induce intestinal epithelial cell apoptosis, a phenomenon associated with acute GVHD. These data suggest that KGF prevents the development of acute lethal GVHD by protecting epithelial cell injury mediated by TNF-alpha, NO, and other potential cytotoxic factors. We noted a moderate reduction in intestinal KGFR mRNA expression in untreated GVH mice on day 8, when IFN-gamma mRNA levels were highest. This reduction in KGFR mRNA levels was not seen in recipients of IFN-gamma gene knockout grafts, suggesting that IFN-gamma may be involved in reducing KGFR mRNA expression in the intestine. A similar reduction in intestinal KGFR mRNA expression was also seen in rHuKGF-treated recipients, suggesting that rHuKGF does not mediate its protective effect by maintaining KGFR at control levels. KGF-treatment also redirected the cytokine response in acute GVH mice from Th1 to a mixed pattern of both Th1 and Th2 cytokines. This was associated with histopathologic changes resembling chronic GVHD.

A novel chordin-like BMP inhibitor, CHL2, expressed preferentially in chondrocytes of developing cartilage and osteoarthritic joint cartilage.

We have identified a novel chordin-like protein, CHL2, which is structurally most homologous to CHL/neuralin/ventroptin. When injected into Xenopus embryos, CHL2 RNA induced a secondary axis. Recombinant CHL2 protein interacted directly with BMPs in a competitive manner to prevent binding to the type I BMP receptor ectodomain, and inhibited BMP-dependent induction of alkaline phosphatase in C2C12 cells. Thus, CHL2 behaves as a secreted BMP-binding inhibitor. In situ hybridization revealed that CHL2 expression is restricted to chondrocytes of various developing joint cartilage surfaces and connective tissues in reproductive organs. Adult mesenchymal progenitor cells expressed CHL2, and its levels decreased during chondrogenic differentiation. Addition of CHL2 protein to a chondrogenic culture system reduced cartilage matrix deposition. Consistently, CHL2 transcripts were weakly detected in normal adult joint cartilage. However, CHL2 expression was upregulated in middle zone chondrocytes in osteoarthritic joint cartilage (where hypertrophic markers are induced). CHL2 depressed chondrocyte mineralization when added during the hypertrophic differentiation of cultured hyaline cartilage particles. Thus, CHL2 may play negative roles in the (re)generation and maturation of articular chondrocytes in the hyaline cartilage of both developing and degenerated joints.

Pharmacodynamics of the type II calcimimetic compound cinacalcet HCl.

Calcimimetic compounds, which activate the parathyroid cell Ca(2+) receptor (CaR) and inhibit parathyroid hormone (PTH) secretion, are under experimental study as a treatment for hyperparathyroidism. This report describes the salient pharmacodynamic properties, using several test systems, of a new calcimimetic compound, cinacalcet HCl. Cinacalcet HCl increased the concentration of cytoplasmic Ca(2+) ([Ca(2+)](i)) in human embryonic kidney 293 cells expressing the human parathyroid CaR. Cinacalcet HCl (EC(50) = 51 nM) in the presence of 0.5 mM extracellular Ca(2+) elicited increases in [Ca(2+)](i) in a dose- and calcium-dependent manner. Similarly, in the presence of 0.5 mM extracellular Ca(2+), cinacalcet HCl (IC(50) = 28 nM) produced a concentration-dependent decrease in PTH secretion from cultured bovine parathyroid cells. Using rat medullary thyroid carcinoma 6-23 cells expressing the CaR, cinacalcet HCl (EC(50) = 34 nM) produced a concentration-dependent increase in calcitonin secretion. In vivo studies in rats demonstrated cinacalcet HCl is orally bioavailable and displays approximately linear pharmacokinetics over the dose range of 1 to 36 mg/kg. Furthermore, this compound suppressed serum PTH and blood-ionized Ca(2+) levels and increased serum calcitonin levels in a dose-dependent manner. Cinacalcet was about 30-fold more potent at lowering serum levels of PTH than it was at increasing serum calcitonin levels. The S-enantiomer of cinacalcet (S-AMG 073) was at least 75-fold less active in these assay systems. The present findings provide compelling evidence that cinacalcet HCl is a potent and stereoselective activator of the parathyroid CaR and, as such, might be beneficial in the treatment of hyperparathyroidism.

In vitro studies with the calcimimetic, cinacalcet HCl, on normal human adult osteoblastic and osteoclastic cells.

This study examined whether the calcium-sensing receptor (CaR) is expressed in normal adult human osteoblastic and osteoclastic cells in culture, and whether the calcimimetic, cinacalcet HCl (AMG 073), potentiates the effects of calcium (via CaR, or some other receptor/mechanism). When mouse or human osteoblastic cells were treated with higher concentrations of calcium (6.6 or 8.6 mM in alpha-MEM/10% FBS) than present in control cultures (1.6 mM), the previously well-documented increase in cell number was demonstrated. Cinacalcet HCl affected cell proliferation of CHO cells transfected with CaR, dose dependently, but had no effect on human or mouse osteoblastic cell proliferation in calcium-containing medium (1.6 or 8.6 mM). To test cinacalcet HCl and calcium on osteoclastic cells, peripheral blood mononuclear cells were cultured in medium containing RANK ligand and M-CSF, supplemented with calcium, and/or cinacalcet HCl. Tartrate-resistant acid phosphatase-positive multinucleated osteoclastic cells on plastic or bone were then counted at 11 and 21 days, respectively. Calcium (greater than 6.0 mM) inhibited osteoclast formation, but cinacalcet HCl (30-1000 nM) had no effect on osteoclastic formation or resorption in the presence of calcium (1.6 or 6.1 mM). RT-PCR did not detect CaR in human, rat, or mouse primary osteoblastic cells and cell lines or osteoclastic cells. In conclusion, these studies indicate that the calcium-induced increase in osteoblastic cell number, and the decrease in formation/function of osteoclastic cells, involves a mechanism or receptor other than CaR. In addition, the calcimimetic agent did not potentiate the effects of calcium on normal adult human bone cells in vitro.

The role of interferon-gamma, nitric oxide and lipopolysaccharide in intestinal graft-versus-host disease developing in F1-hybrid mice.

(C57BL/6 x DBA/2)F1-hybrid mice injected with lymphoid cells from wild-type, C57BL/6 donors develop acute, lethal graft-versus-host disease (GVHD) in which the intestine is a major target. In its destructive phase intestinal GVHD is characterized by apoptosis of intestinal crypt epithelial cells and the development of endotoxaemia. Injection of as little as 10 microg endotoxin is lethal in mice with acute GVHD, and associated with the release of large amounts of tumour necrosis factor-alpha (TNF-alpha) into the serum. To explore the role of interferon-gamma (IFN-gamma) in the pathogenesis of intestinal GVHD we used IFN-gamma gene knockout (gko) mice as donors. Recipients of grafts from these donors did not develop intestinal GVHD and, unlike recipients of wild-type grafts, did not die when injected with lipopolysaccharide (LPS). We also found that injection 10 microg LPS into recipients of wild-type grafts induced apoptosis of intestinal epithelial crypt cells and was associated with a burst of nitric oxide production in the intestine. Administration of N(omega)nitro L-arginine methyl ester blocked this response. In contrast, LPS did not induce either intestinal epithelial cell apoptosis or increased nitric oxide production in recipients of IFN-gamma gko grafts. These findings indicate that donor-derived IFN-gamma is instrumental for the development of intestinal GVHD. In a previous study we showed that recipients of IFN-gamma gko grafts develop high levels of LPS-induced TNF-alpha release. When our current data are viewed in the context of this observation, they suggest that intestinal epithelial cell apoptosis in the parent-->F1-hybrid model of acute GVHD is mediated primarily by nitric oxide rather than TNF-alpha, and that this depends on donor-derived IFN-gamma.

SKIP3, a novel Drosophila tribbles ortholog, is overexpressed in human tumors and is regulated by hypoxia.

Regions of hypoxia are a hallmark of solid tumors. Tumor cells modulate the regulation of specific genes allowing adaptation and survival in the harsh hypoxic environment. We have identified SKIP3, a novel human kinase-like gene, which is overexpressed in multiple human tumors and is regulated by hypoxia. SKIP3 is an ortholog of the Drosophila tribbles, rat NIPK, dog C5FW, and human C8FW genes. Drosophila tribbles is involved in slowing cell-cycle progression during Drosophila development, but little is known regarding the function or tissue distribution of the vertebrate orthologs. We show that the normal tissue expression of SKIP3 is confined to human liver, while multiple primary human lung, colon, and breast tumors express high levels of SKIP3 transcript. Endogenous SKIP3 protein accumulates within 48 h under hypoxic growth conditions in HT-29 and PC-3 cells, with upregulation of the SKIP3 mRNA transcript by 72 h. We identified activating transcription factor 4 (ATF4) as a SKIP3-binding partner using the yeast-two-hybrid assay. Coexpression of SKIP3 and ATF4 showed that SKIP3 is associated with the proteolysis of ATF4, which can be blocked using a proteosome inhibitor. These results indicate that SKIP3 may be an important participant in tumor cell growth.

Inhibition of interleukin-1 but not tumor necrosis factor suppresses neovascularization in rat models of corneal angiogenesis and adjuvant arthritis.

OBJECTIVE: To assess the capacities of the cytokine inhibitors interleukin-1 receptor antagonist (IL-1Ra; anakinra) and PEGylated soluble tumor necrosis factor receptor I (PEG sTNFRI; pegsunercept) to suppress neovascularization. METHODS: A corneal angiogenesis assay was performed by implanting nylon discs impregnated with an angiogenic stimulator (basic fibroblast growth factor or vascular endothelial growth factor) into one cornea of female Sprague-Dawley rats. Animals were treated with IL-1Ra or PEG sTNFRI for 7 days, after which new vessels were quantified. In a parallel study, male Lewis rats with mycobacteria-induced adjuvant-induced arthritis were treated with IL-1Ra or PEG sTNFRI for 7 days beginning at disease onset, after which scores for inflammation and bone erosion as well as capillary counts were acquired from sections of arthritic hind paws. RESULTS: Treatment with IL-1Ra yielded a dose-dependent reduction in growth factor-induced corneal angiogenesis, while PEG sTNFRI did not. IL-1Ra, but not PEG sTNFRI, significantly reduced the number of capillaries in arthritic paws, even though both anticytokines reduced inflammation and bone erosion to a similar degree. CONCLUSION: These data support a major role for IL-1, but not TNFalpha, in angiogenesis and suggest that an additional antiarthritic mechanism afforded by IL-1 inhibitors, but not anti-TNF agents, is the suppression of the angiogenic component of pannus.

Transgenic overexpression of human IL-17E results in eosinophilia, B-lymphocyte hyperplasia, and altered antibody production.

We have identified and cloned a novel human cytokine with homology to cytokines of the interleukin-17 (IL-17) family, which we have termed human IL-17E (hIL-17E). With the identification of several IL-17 family members, it is critical to understand the in vivo function of these molecules. We have generated transgenic mice overexpressing hIL-17E using an apolipoprotein E (ApoE) hepatic promoter. These mice displayed changes in the peripheral blood, particularly, a 3-fold increase in total leukocytes consisting of increases in eosinophils, lymphocytes, and neutrophils. Splenomegaly and lymphoadenopathy were predominant and included marked eosinophil infiltrates and lymphoid hyperplasia. CCR3(+) eosinophils increased in the blood and lymph nodes of the transgenic mice by 50- and 300-fold, respectively. Eosinophils also increased 8- to 18-fold in the bone marrow and spleen, respectively. In the bone marrow, most of the eosinophils had an immature appearance. CD19(+) B cells increased 2- to 5-fold in the peripheral blood, 2-fold in the spleen, and 10-fold in the lymph nodes of transgenic mice, whereas CD4(+) T lymphocytes increased 2-fold in both blood and spleen. High serum levels of the cytokines IL-2, IL-4, IL-5, granulocyte colony-stimulating factor, eotaxin, and interferon gamma were observed. Consistent with B-lymphocyte increases, serum immunoglobulin (Ig) M, IgG, and IgE were significantly elevated. Antigenic challenge of the transgenic mice with keyhole limpet hemocyanin (KLH) resulted in a decrease in anti-KLH IgG accompanied by increases of anti-KLH IgA and IgE. In situ hybridization of transgenic tissues revealed that IL-17Rh1 (IL-17BR/Evi27), a receptor that binds IL-17E, is up-regulated. Taken together, these data indicate that IL-17E regulates hematopoietic and immune functions, stimulating the development of eosinophils and B lymphocytes. The fact that hIL-17E overexpression results in high levels of circulating eosinophils, IL-4, IL-5, eotaxin, and IgE suggests that IL-17E may be a proinflammatory cytokine favoring Th2-type immune responses.

rHuKGF ameliorates symptoms in DSS and CD4(+)CD45RB(Hi) T cell transfer mouse models of inflammatory bowel disease.

There is an acute need for effective therapy for inflammatory bowel disease (IBD), particularly at the level of repair of the damaged epithelium. We evaluated the efficacy of recombinant human keratinocyte growth factor (rHuKGF) in both the dextran sodium sulfate (DSS) and the CD4(+)CD45RB(Hi) T cell transfer models of IBD. Disease was induced either by the ad libitum administration to normal mice of 4% DSS in the drinking water or by the injection of 4 x 10(5) CD4(+)CD45RB(Hi) T cells into immunodeficient scid/scid mice. rHuKGF was administered by subcutaneous injection at doses of 1.0 or 3.0 mg/kg in both preventative and therapeutic regimens during both studies. rHuKGF significantly improved survival and body weight loss in the DSS model in both preventative and therapeutic dosing regimens. It also improved diarrhea, hematochezia, and hematological parameters, as well as large intestine histopathology. In the T cell transfer model, rHuKGF improved body weight loss, diarrhea, and levels of serum amyloid A, as well as large intestine histopathology. In both models of IBD, the colonic levels of intestinal trefoil factor (ITF) were elevated by the disease state and further elevated by treatment with rHuKGF. These data suggest that rHuKGF may prove useful in the clinical management of IBD and its effects are likely mediated by its ability to locally increase the levels of ITF.