Multiregion whole-exome sequencing of matched primary and metastatic tumors revealed genomic heterogeneity and suggested polyclonal seeding in colorectal cancer metastasis.

BACKGROUND: Distant metastasis accounts for 90% of deaths from colorectal cancer (CRC). Genomic heterogeneity has been reported in various solid malignancies, but remains largely under-explored in metastatic CRC tumors, especially in primary to metastatic tumor evolution. PATIENTS AND METHODS: We conducted high-depth whole-exome sequencing in multiple regions of matched primary and metastatic CRC tumors. Using a total of 28 tumor, normal, and lymph node tissues, we analyzed inter- and intra-individual heterogeneity, inferred the tumor subclonal architectures, and depicted the subclonal evolutionary routes from primary to metastatic tumors. RESULTS: CRC has significant inter-individual but relatively limited intra-individual heterogeneity. Genomic landscapes were more similar within primary, metastatic, or lymph node tumors than across these types. Metastatic tumors exhibited less intratumor heterogeneity than primary tumors, indicating that single-region sequencing may be adequate to identify important metastasis mutations to guide treatment. Remarkably, all metastatic tumors inherited multiple genetically distinct subclones from primary tumors, supporting a possible polyclonal seeding mechanism for metastasis. Analysis of one patient with the trio samples of primary, metastatic, and lymph node tumors supported a mechanism of synchronous parallel dissemination from the primary to metastatic tumors that was not mediated through lymph nodes. CONCLUSIONS: In CRC, metastatic tumors have different but less heterogeneous genomic landscapes than primary tumors. It is possible that CRC metastasis is, at least partly, mediated through a polyclonal seeding mechanism. These findings demonstrated the rationale and feasibility for identifying and targeting primary tumor-derived metastasis-potent subclones for the prediction, prevention, and treatment of CRC metastasis.

Prognostic significance of high-grade dysplasia in colorectal adenomas.

AIM: Colonoscopy to detect and remove polyps has contributed to a reduction in colorectal carcinoma. Three-year follow up is recommended for patients considered to be at high risk (at least three adenomas, adenoma ≥ 1 cm, villous or high-grade features). Our study focused on patients diagnosed with high-grade dysplasia with regard to initial management and follow up. METHOD: A search of patients who had had endoscopic removal of a high-grade adenoma was carried out. Patients with the following were excluded: follow up of < 1 year, polyposis syndromes, prior colon cancer and a diagnosis of adenocarcinoma within 6 months following initial diagnosis. RESULTS: Eighty-three patients treated between 1999 and 2007 for high-grade dysplasia (HGD) in a colorectal adenoma were identified. Over a median follow-up period of 4 years, 53 (64%) developed further adenomatous polyps. Among these, 7% had an adenoma with HGD or an adenocarcinoma. In all these patients, the initial high-grade adenoma was > 1 cm in diameter. Initial follow-up colonoscopy was performed on average 7 months following the initial diagnosis. Ten per cent of patients underwent prophylactic segmental resection, and 6% received argon laser therapy. CONCLUSION: The study demonstrates that patients who have a colorectal adenoma > 1 cm with HGD may be at high risk of developing further adenomas with HGD or carcinoma. Close follow up is warranted.

Loss of FHIT expression in gastric carcinoma.

Loss of heterozygosity involving the short arm of chromosome 3 has been reported in gastric and other human tumors. We have cloned and mapped a candidate tumor suppressor gene, FHIT (fragile histidine triad), to this chromosomal region (3p14.2). To investigate the role of FHIT gene alterations in the development of gastric carcinoma, we examined 8 gastric carcinoma-derived cell lines and 32 primary adenocarcinoma samples by Southern blot analysis. We also analyzed the integrity of FHIT transcripts by reverse transcription-PCR. The occurrence of alterations in the FHIT gene and its transcript correlated with the absence of Fhit protein expression by immunoblot analysis in the cancer cell lines. Four of eight cell lines showed deletion or rearrangement within the FHIT gene, together with the absence of the wild-type transcript and the Fhit protein. Among the primary gastric carcinomas, rearrangement of the FHIT gene and/or aberrant reverse transcription-PCR products were detected in 17 of 32 (53%) tumors, and 20 of 30 (67%) samples exhibited an absence of Fhit protein expression. Gastric cancer is thought to develop from carcinogenic exposure, possibly explaining the high frequency of abnormalities in the FHIT gene, a fragile locus exhibiting susceptibility to carcinogen-induced alterations. The consequent absence or reduction of Fhit protein expression is consistent with the proposal that the FHIT gene is a preferential target of environmental carcinogens and that FHIT inactivation plays a role in the development of gastric cancer.

Endosonographic and color Doppler flow imaging alterations observed within irradiate rectal cancer.

PURPOSE: To correlate the endosonographic and color Doppler flow imaging alterations observed in irradiated rectal cancers with the pathologic features of radiation response, and to evaluate the potential impact of altered blood flow on the integrity of the surgical anastomosis. METHODS AND MATERIALS: Endosonography with color and pulsed wave Doppler was performed on 20 rectal cancer masses before and after high dose preoperative radiation (XRT). Pre- and post-XRT observations included comparing alterations in tumor size, sonographic echotexture, color Doppler flow, and pulsatility indices. Comparisons were made with pathologic findings in the irradiated specimens and with the incidence of anastomotic failure. RESULTS: Compared to pre-XRT observations, irradiated rectal cancers decreased in size and became either mixed in echogenicity with less apparent color Doppler flow (16 of 20) or unchanged in color Doppler flow and echotexture (4 of 20). Those with less flow (16 of 20) were imaged later (mean = 90.2 +/- 12.1 days) than those without change in color Doppler flow (mean = 21.7 +/- 2.7 days). Pathologically, the group of four without change in color Doppler signal had features of acute inflammation which were not observed in 16 of 20 imaged later. Based on pulsatility index measurements, both high and low resistance vessels were detected and confirmed by immunohistochemical staining, and features of postradiation obliterative vasculitis were observed. Only one primary anastomosis in 14 patients with decreased flow failed. CONCLUSIONS: The sonographic and color Doppler flow imaging alterations observed within irradiated rectal cancer correlated with changes of postradiation obliterative vasculitis. The apparent diminished local blood flow within high and low resistance vessels post-XRT did not result in an increased incidence of anastomotic failures.

Pathologic features of Hashimoto's-associated papillary thyroid carcinomas.

Some investigators have found an increased incidence of papillary carcinoma (PC) of the thyroid in patients with Hashimoto's (autoimmune) thyroiditis (HT), which raises the possibility that there may be more than an incidental association between these 2 diseases. In this study, we analyzed the pathology of Hashimoto's-associated thyroid carcinomas to see if these tumors showed any distinctive features. The possible significance of solid cellular nodules as preneoplastic lesions in patients with HT was investigated. A review of all the cases of HT during a 16-year period yielded 30 PC and 3 follicular carcinomas (FC). Within the PC there were 7 (23%) follicular variants. Twenty (67%) of the PC showed various degree of intratumoral fibrosis, ranging from thick fibrous septa separating tumor nodules to almost complete obliteration of the tumor by the fibrosis, with only microscopic residual tumor nests. In most of the cases, the desmoplastic response within the tumors was of the fibromatosis-like type with dense hyalinized collagen and bland-appearing spindle cells. All the tumors, independently of the degree of fibrosis, showed the nuclear features of PC. No correlation was found between the degree of fibrosis in the tumors and the thyroid gland outside the tumors. There were tumors with marked fibrosis without fibrosis outside the tumors. Four cases of PC (13%) showed a growth pattern characterized by cystic spaces with thick hyalinized walls and focal papillary hyperplasia lined by flat and cuboidal epithelium, reminiscent of a vascular neoplasm. There were 4 atypical solid microscopic nodules with confluent cellularity; 2 of them associated with a PC and the other 2 with diffuse HT without PC. These nodules were composed of cells with clear nuclei and occasional grooves without nuclear pseudoinclusions. By immunohistochemistry, 2 of 3 nodules showed cytoplasmic reactivity for cytokeratin 19, and 2 of 3 nodules were positive for the RET/PTC (rearranged during transfection, papillary thyroid carcinoma) antibody. In summary, HT-associated PC may frequently display prominent stromal desmoplasia and a pseudovascular pattern, both of which can present diagnostic difficulties if the cytologic features of PC are not recognized because of the marked obliteration of the tumor by the fibrosis. Atypical nodules may represent a precursor lesion of PC in patients with HT.

Immunohistochemical localization of p21(WAF1/CIP1) in normal, hyperplastic, and neoplastic uterine tissues.

p21WAF1/CIP1 is a nuclear protein that binds to cyclin-dependent kinase complexes (CDKs) and inhibits the activity of multiple kinases. These CDKs are involved in the regulation of cell cycle progression at several checkpoints. In this study, the authors have analyzed by immunohistochemistry the expression of p21WAF1/CIP1 in normal uterine tissues, 12 endometrial hyperplasias, 17 endocervical adenocarcinomas, and 31 endometrial adenocarcinomas. In addition, a group of 10 leiomyomas and 10 uterine leiomyosarcomas were also stained. To evaluate cell proliferation, the monoclonal antibody Ki-67 was used in all of the available cases. Terminally differentiated epithelial endocervical and endometrial cells showed variable expression of p21WAF1/CIP1, whereas the endometrial hyperplasias, and endocervical and endometrial adenocarcinomas showed decreased expression or were negative. All of the cases of cervical squamous dysplasia were positive. Normal smooth muscle cells and 50% of leiomyomas were negative, whereas all leiomyosarcomas showed expression of p21WAF1/CIP1. These results indicate that p21WAF1/CIP1 contributes to differentiation in normal endometrial and endocervical glands. The decreased expression of p21WAF1/CIP1 in endometrial hyperplasias and carcinomas may be important in the process of neoplastic transformation. The role of certain CDK inhibitors, such as p21WAF1/CIP1, is different in epithelial and mesenchymal tumorigenesis in the uterus.

Colorectal carcinomas with high microsatellite instability: defining a distinct immunologic and molecular entity with respect to prognostic markers.

Molecular analysis of hereditary nonpolyposis colorectal carcinomas (HNPCC) has identified DNA mismatch repair deficiencies with resulting microsatellite instability (MSI) as a pathway of carcinogenesis that appears to be relevant for prognosis, treatment, and possibly prevention. In this study, expression of cell cycle proteins and other known prognostic markers is correlated with the microsatellite status of colorectal cancers (CRC). One hundred consecutive cases from the CRC Registry at Thomas Jefferson University were analyzed for MSI. Immunohistochemistry was performed for the mismatch repair proteins hMLH1 and hMSH2, tumor suppressor p53, apoptosis inhibitor bcl-2, cell cycle proteins p21(WAF1/CIP1), and p27 and the proliferation markers Ki-67 and topoisomerase II. High MSI (MSI-H) is significantly correlated with loss of either hMLH1 or hMSH2, presence of bcl-2, and absence of p53. p21(WAF1/CIP1) is positive in all tumors with MSI-H. Previous findings of a lower proliferation rate were confirmed with a topoisomerase II stain. Microsatellite stable (MSS) tumors generally express both MSH2 and MLH1. Other highly significant differences are positive p53 in 56% of MSS cases and negative bcl-2 in 98% of MSS cases. p27 expression is found in approximately 50% of all CRCs irrespective of the microsatellite status. MSI-H tumors follow the mutator pathway, with loss of expression of one mismatch repair protein, wild-type p53, lower proliferation, and positivity for p21(WAF1/CIP1). MSS tumors follow the suppressor pathway, characterized by p53 overexpression, higher proliferation, and absence of bcl-2 expression; p21(WAF1/CIP1) expression can be variable. These data provide a molecular basis for the clinical observation that patients with HNPCC appear to have a more favorable prognosis. HUM PATHOL 31:1506-1514.

The role of p53, p21WAF1/C1PI, and bcl-2 in radioresistant colorectal carcinoma.

Genetic alterations in the p53 tumor suppressor gene are common in human colorectal cancers, occurring in approximately 70% of tumors. In vitro studies have shown that wild-type p53 is involved in controlling cell cycle checkpoint functions and apoptosis involved in the cytotoxic response induced by ionizing radiation and several anticancer chemotherapeutic agents. Wild-type p53 protein can transcriptionally activate the WAF gene, which encodes a cyclin-dependent kinase inhibitory protein, p21WAF1/C1PI protein, and transcriptionally repress the bcl-2 gene, which encodes an inhibitor of apoptosis. To learn more about the in vivo relationship between p53 protein and the expression of p21WAF1/C1PI and bcl-2 proteins in human colorectal cancers treated with radiation therapy, we examined the expression of these proteins by immunohistochemistry in pre-irradiated biopsy specimens and surgical specimens with residual tumor of 27 patients with colorectal carcinoma. Cell proliferation was measured using Ki-67 expression in the tumor cells. The p53 protein was not detected in normal colorectal mucosa, but it was expressed in 21 of 27 (78%) of pre-irradiated tumor samples and in 19 of 27 (70%) of post-irradiated tumors. Expression of the bcl-2 protein in normal colorectal mucosa was confined to the basal epithelial cells of the crypts. Diffuse bcl-2 staining was detected in tumor cells in 13 of 27 (48%) of pre-irradiated samples and in 14 of 27 (52%) of post-irradiated samples. p21WAF1/C1PI expression was detected in 14 of 27 (52%) of pre-irradiated samples but only in 7 of 27 (26%) of post-irradiated samples. No inverse relationship between expression of p53 protein and abnormal bcl-2 expression was apparent. p21WAF1/C1PI was expressed in most nonproliferating Ki-67-negative epithelial cells at the apical tips of the crypts in normal colorectal mucosa, but not in proliferating Ki-67-positive cells of adjacent adenomatous mucosa. An inverse relationship between Ki-67 and p21WAF1/C1PI expression was observed in normal colorectal mucosa and adjacent adenomatous mucosa. After radiation therapy, p53 protein accumulation did not change among residual tumors in 18 cases (three of which were initially negative and remained negative); in four cases there was a significant increase, and five cases had a substantial decrease of p53 expression. Aberrant bcl-2 expression is not correlated with expression of p53 and does not increase significantly in post-irradiated tumor cells. p21WAF1/C1PI expression is markedly reduced in tumor cells that survive radiation therapy.

Overexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline ovarian tumors.

Ovarian epithelial tumors are classically divided into benign, malignant, and borderline or of low malignant potential. It is controversial whether this last group of tumors should be considered benign or malignant. Expression of cell cycle markers has recently been linked to tumor behavior and response to treatment. It has been shown that one of the pathways through which the p53 gene controls the cell cycle is by transactivating p21WAF1/CIP1, a cyclin-dependent kinase (cdk) inhibitor. By inhibiting cdks, p21WAF1/CIP1 blocks the G-1 to S-phase transition in the cell cycle. p53 can be regulated by MDM2 (murine double minute-2) through direct inactivation or promotion of its cytoplasmic degradation. In an attempt to investigate the cell cycle checkpoint mechanisms of these tumors, we studied the expression of p53, Ki-67, MDM2, and p21WAF1/CIP1 by immunohistochemistry. We analyzed the expression of these proteins in 19 cystadenomas (8 serous and 11 mucinous), 40 borderline tumors (31 serous and 9 mucinous), and 18 serous carcinomas of the ovary. p21WAF1/CIP1 was expressed in 7 of 19 (37%) benign cystadenomas, 32 of 40 (80%) borderline tumors (93.5% of serous and 33% of mucinous), and in 9 of 18 (50%) serous carcinomas. Ki-67 was only weakly expressed in 8 of 19 (42%) benign cystadenomas, all borderline tumors showed Ki-67 staining in less than 50% of the cells, and 55% of serous carcinomas stained in more than 50% of tumor cells. p53 was absent in all but 1 of the cystadenomas, was expressed in 9 of 40 (22.5%) borderline tumors (25.8% of serous and 11% of mucinous), and in 10 of 18 (55%) carcinomas. All 11 implants of serous borderline tumors expressed p21WAF1/CIP1. Most serous borderline tumors expressed higher levels of MDM2 compared with the benign cystadenomas and carcinomas. Four of the serous borderline implants (40%) expressed MDM2. Coexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline tumors of the ovary and their implants, which suggests that these cell cycle control proteins are important in these tumors and may be related to tumor progression. Low expression of p53 protein in serous borderline tumors might be in part mediated by MDM2. This suggests that the p53 pathway is intact in most of these tumors, in contrast with carcinomas, in which high expression of p53 has been related to mutations of this gene.

Reciprocity between membranous and nuclear expression of beta-catenin in colorectal tumours.

beta-Catenin has a central role not only in linking the cadherin-mediated cell adhesion system but also in the intercellular signalling pathway. To investigate alterations of beta-catenin in the development of colorectal carcinoma, the pattern of beta-catenin expression was studied using immunohistochemistry in 74 sporadic colorectal adenomas, in histologically normal mucosa adjacent to 65 of these adenomas, and in 52 carcinomas arising in adenomas. All normal epithelia displayed cell boundary staining for beta-catenin. Adenomas and carcinomas showed varying degrees of membranous staining. However, some tumours also showed nuclear staining of beta-catenin protein. Decreased membranous and increased nuclear beta-catenin staining were associated with increasing degrees of dysplasia in adenomas (P < 0.005, P < 0.05, respectively). Carcinomas manifested significantly reduced membranous, but enhanced nuclear beta-catenin expression compared with their associated adenomas (P < 0.001, P < 0.005, respectively). An inverse correlation was found between decreased membranous and increased nuclear staining of beta-catenin in both adenomas and carcinomas (P < 0.025, P < 0.05, respectively). The data confirm that reduced membranous and increased nuclear expression of beta-catenin is associated with the progression of colorectal adenomas to carcinomas. Our results also suggest that decreased membranous expression of beta-catenin may result from aberrant localisation of the protein in the cell nucleus.

Cytogenetic findings in a breast stromal sarcoma. Application of fluorescence in situ hybridization to characterize the breakpoint regions in an 11;19 translocation.

Cytogenetic analysis of a stromal breast sarcoma revealed a complex karyotype that included a reciprocal 11;19 translocation, along with multiple numerical changes, deletions, and other unbalanced structural rearrangements. Karyotypic abnormalities have not been reported previously in this rare neoplasm that arises from mesenchymal breast tissue, and the t(11;19) is of interest because various types of sarcoma are characterized by specific reciprocal translocations. Because of the pericentric nature of the breakpoints on chromosomes 11 and 19 in the t(11;19), classical cytogenetic banding could not reveal the centromeric origin of the translocation derivatives. Using nonisotopic in situ hybridization with chromosome 11 and 19 alpha-satellite probes, the centromere of each derivative chromosome was determined, and the rearrangement was interpreted as a balanced translocation, t(11;19)(q12 or q13.1;p12 or p13.1). This abnormality has not been described previously in any breast tumor.

Reduced expression of molecules of the cadherin/catenin complex in the transition from colorectal adenoma to carcinoma.

E-cadherin is crucial to the intercellular adherens junctions which are involved in the organisation and maintenance of epithelial structure and suppression of tumour invasion. E-cadherin is associated with the actin cytoskeleton via cytoplasmic proteins, including alpha-, beta- and gamma-catenins, which together form the cadherin/catenin complex. To evaluate changes of the molecules of the cadherin/catenin complex in colorectal carcinogenesis, seventy-four sporadic adenomas, samples of histologically normal epithelium adjacent to 65 adenomas, and 52 carcinomas arising in adenomas were investigated by immunohistochemistry. All normal epithelial cells showed a uniform membranous staining pattern for E-cadherin, alpha-, beta-, and gamma-catenin. Decreased expression of all 4 proteins occurred in parallel in adenomas and carcinomas (in all cases, p < 10(-5). Decreased expression of the cadherin/catenin complex in adenomas was associated with increasing severity of dysplasia (p < 0.001, for E-cadherin, alpha-, and gamma-catenin, p < 0.005 for beta-catenin). Carcinomas displayed significantly reduced expression of the cadherin/catenin complex compared with their associated adenomas (all p < 0.001). The results directly confirm that colorectal tumour progression and invasion is associated with disruption of the cadherin/catenin complex and suggest that the genetic changes and transcriptional modulation of catenins underlying this progression may affect all members of the complex.

A comparative analysis of Ki-67, p53, and p21WAF1CIP1 expression in tamoxifen associated endometrial carcinomas.

OBJECTIVE: To determine clinicopathologic parameters, expression of proliferation markers, and immunohistochemical oncogene expression in endometrial cancers in patients with a history of breast cancer with and without tamoxifen use. METHODS: Thirty endometrial carcinoma specimens were examined from patients with a previous history of breast cancer. Patients who had taken tamoxifen (15) were compared to non-users (15). Immunohistochemical staining was performed for p53, Ki-67, and p21WAF1/CIP1, overexpression was defined as greater than 10% positivity. RESULTS: Patient populations were statistically similar. P53 was overexpressed in 73% of tamoxifen users compared to 53% of non-users. Ki-67 was overexpressed in over 90% of user and non-user specimens. p21WAF1/CIP1 was overexpressed in 33% of users and 47% of non-users. Tamoxifen users had shorter time to diagnosis of endometrial cancer than non-users. CONCLUSIONS: In this small study, tamoxifen associated tumors expressed p53 more frequently than non-users, while the opposite was observed with p21WAF1/CIP1. This suggests that p53 mutations might play a role in development of tamoxifen associated tumors.

Endometrial hyperplasia and the risk of carcinoma.

Recent reports suggest that atypical endometrial hyperplasia diagnosed by biopsy or curettage is accompanied by a higher than expected risk of coexistent invasive cancer. In order to test this hypothesis we reviewed the pathology and clinical history of all patients at our institution who underwent hysterectomy for endometrial hyperplasia with or without cytologic atypia. We found 24 patients of 45 with a preoperative diagnosis of hyperplasia with cytologic atypia, and 21 with simple or complex hyperplasia without atypia. No cancers were found at surgery in the latter group nor were any significant historical differences found between the two groups. Of the patients with atypia, 12/24 (50%) had an endometrial carcinoma and nine patients (37.5%) were stage IB or greater. This is a significantly greater risk than previously reported in the literature. Endometrial hyperplasia with cytologic atypia may carry a higher risk of coexistent invasive endometrial carcinoma than previously believed. Methods to identify those patients at highest risk should be determined.

Expression of p21WAF1/CIP1 in soft tissue sarcomas: a comparative immunohistochemical study with p53 and Ki-67.

The p53 gene controls the cell cycle by transactivating p21WAF1/CIP1, a cyclin dependent kinase (cdk) inhibitor. By inhibiting cdks, p21WAF1/CIP1 regulates the cell cycle by blocking the G1 to S phase transition. In this study, we analyzed the immunohistochemical expression of p21WAF1/CIP1 in 66 soft tissue sarcomas and its relationship to p53 and the cell cycle proliferation antigen Ki-67. Expression of p21WAF1/CIP1, was detected in 76% of the tumors and p53 in 26%. All malignant schwannomas, synovial sarcomas, leiomyosarcomas and gastrointestinal stromal tumors expressed p21WAF1/CIP1. The majority of angiosarcomas, dermatofibrosarcomas, and fibrosarcomas showed low expression or were negative for p21WAF1/CIP1. Ewing's sarcomas, liposarcomas, and malignant fibrous histiocytomas were heterogeneous in their expression of p21WAF1/CIP1. Combining p53 and p21WAF1/CIP1 staining, the following four patterns were observed: 23% of the tumors showed the p53+/p21+ pattern; 53% showed the p53-/p21+ pattern; 3% showed the p53+/p21- pattern and 21% were negative for both p53 and p21WAF1/CIP1. There was no correlation between Ki-67 and p21WAF1/CIP1 or p53 staining. Our results show that soft tissue sarcomas, independent of their histologic subtype, frequently express p21WAF1/CIP1 which is probably important in their tumorigenesis. Additionally, p21WAF1/CIP1 may play a role in determining the efficacy of various cell cycle-directed therapies in soft tissue sarcomas.

Treatment of inflammatory bowel disease in a rodent model with the intestinal growth factor glucagon-like peptide-2.

BACKGROUND/PURPOSE: Microinjection of a Fisher (F344) rat zygote with human HLA-B27 and beta2-microglobulin genes induces spontaneous chronic gastrointestinal (GI) inflammation similar to lesions seen in patients with inflammatory bowel disease (IBD). This study was designed to evaluate the potential therapeutic benefit of GLP-2, an intestinal growth factor, in this transgenic rat model of IBD. METHODS: Five F344 (control) and 10 HLA-B27 (on a F344 background) rats at 25 weeks of age were used. Rats were divided into the following 3 groups: group 1, F344 rats, no treatment (n = 5); group 2, HLA-B27, no treatment (n = 5); and group 3, HLA-B27, treated with a 14-day systemic infusion (via the jugular vein) of GLP-2 at 50 microg/kg/d (n = 5). After infusion, all rats underwent laparotomy, and the intestine from the ligament of Treitz to the rectum was harvested. Total mucosal damage (percent surface area) was measured using image analysis software (Sigmascan 2.0). Microscopic analysis was performed by a blinded reviewer and scored as follows: 0, no inflammation; 1, mild inflammation; 2, moderate inflammation; and 3, severe inflammation. Colonic mucosal total RNA was assayed for tumor necrosis factor alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), internal standard, mRNA by reverse transcriptase polymerase chain reaction. Statistical analysis was performed using analysis of variance (ANOVA) and expressed as mean +/- SEM. RESULTS: Normal F344 rats did not show evidence of gross or histological lesions in the small or large intestine. GLP-2 reduced total mucosal damage from 9.0% +/- 0.7% in group 2 to 0.9% +/- 0.5% in group 3 (P < .01). The histological lesion score was reduced from 7.0 +/- 0.6 in group 2 to 4.4 +/- 0.8 in group 3 (P < .01). Furthermore, GLP-2 reduced the mean band intensity (MBI) of TNF-alpha (0.4 +/- 0.04 in group 2 to 0 in group 3, P < .01) and IFN-gamma (0.3 +/- 0.02 in group 2 to 0 in group 3, P < .01). CONCLUSIONS: These data show for the first time that GLP-2 significantly reduces gross (90% decrease) and histological (40% decrease) lesions in this rat model of IBD. This is further supported by a significant decrease in gene expression of the inflammatory mediators TNF-alpha (100% decrease) and IFN-gamma (100% decrease). These data suggest a potential therapeutic role for GLP-2 in IBD.