Differential profile of protumor immunological factors between the tumor site and the tumor-free site - predictive potential of IL-8 and COX2 for colorectal cancer and metastasis.

To study the role of host immune surveillance in the initiation and progression of colorectal cancer (CRC), a set of protumor immunological factors was determined by quantitative real-time PCR (q-PCR) between the primary tumor and the adjacent tumor-free site tissues in 63 patients with colorectal neoplasms. Results showed that expression levels of interleukin (IL)-1ß, IL-6, IL-8, IL-17A, IL-23, and cyclooxygenase 2 (COX2) mRNAs, except transforming growth factor beta (TGFß), in adenoma tissues were significantly higher than that in relative adjacent tissues. Difference of immunological factor levels between adenoma and adjacent tissues (Δ values) was in an order of ΔIL-8 > ΔIL-6 > ΔIL-17A > ΔIL-1ß > ΔCOX2 > ΔIL-23; Analysis showed that the value of ΔCOX2 correlated to the grade of dysplastic degree in patients with adenoma. Notably, levels of all these immunological factors in CRC tissues were continuously increased, the order of values of Δ immunological factors was ΔIL-8 > ΔCOX2 > ΔIL-6 > ΔIL-1ß > ΔIL-17A > ΔIL-23 > ΔTGFß. Further analysis revealed that increased value of Δ IL-1ß was associated with advanced TNM stage, a higher value of Δ COX2 tended to predicate a deeper degree of tumor invasion; and higher values of Δ IL-1ß, IL-6 and COX2 closely correlated to lymph node metastasis in patients with CRC. In addition, the ratio of ΔIL-8/ΔTGFß was most obvious changed factor and associated with node metastasis in patients with CRC. Therefore, we concluded that the difference of protumor immunological factor levels between the primary tumor site and tumor-free site along the adenoma-carcinoma sequence reflects the change of protumor/antitumor force balance, which is associated with CRC initiation and invasion.

Occurrences and phenotypes of RIPK3-positive gastric cells in Helicobacter pylori infected gastritis and atrophic lesions.

BACKGROUND: Research evidences suggest that diverse forms of programmed cell death (PCD) are involved in the helicobacter pylori (H. pylori)-induced gastric inflammation and disorders. AIMS: To characterize occurrences and phenotypes of necroptosis in gastric cells in H. pylori infected gastritis and atrophic specimens. METHODS: Occurrences and phenotypes of necroptosis in gastric cells were immunohistochemically characterized with receptor-interacting protein kinase 3 (RIPK3) antibody in both human H. pylori infected gastric gastritis, atrophic specimens, and transgenic mice. RESULTS: Increased populations of RIPK3-positive cells were observed in both gastric glands and lamina propria in H. pylori infected human oxyntic gastritis and atrophic specimens. Phenotypic analysis revealed that many RIPK3-positive cells were H + K+ ATPase-positive parietal cells in the gastric glands and were predominantly CD3-positive T lymphocytes, CD68-positive macrophages, and SMA-alpha-positive stromal cells in the lamina propria. Furthermore, we found an increased expression of RIPK3-positive gastric glandular cells along with the histological process of hyperplasia-atrophy-dysplasia progression in hypergastrinemic INS-GAS mice. CONCLUSIONS: An increased population of RIPK3-positive cells was observed in several types of gastric cells, future studies that define the effects and mechanisms of PCD implicated in the development of H. pylori induced gastric disorders are needed.

ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis.

ST2 (also known as IL1RL1) is the critical functional receptor for interleukin (IL)-33 in stimulating regulatory T cell (Treg) expansion and function in physiological and pathological conditions. We examined the correlation between ST2 cell expression and FoxP3 positive Tregs in both colorectal adenoma and cancer (CRC) microenvironment by real-time PCR, immunohistochemistry (IHC) and double immunofluorescences. The clinicopathological and prognostic significance of cellular ST2-positive cells and FoxP3-positive Tregs in patients with adenoma and CRC were evaluated. Real-time PCR results revealed increased expression levels of ST2 and FoxP3 mRNAs in both adenoma and CRC tissues as compared with control tissues. IHC analysis confirmed increased densities of ST2-positive cells in both the adenoma/CRC epithelium and stroma, which show a close positive linear association with the densities of FoxP3-positive Tregs in respective compartments. Pathological feature analysis showed that densities of ST2-positive cells in the tumor stroma were notably associated with degree of dysplastic grading in patients with adenoma, and disease stages and lymph node metastasis in patients with CRC. Kaplan-Meier survival curves suggested that CRC patients with high densities of ST2-positive cells in the stroma tend to have a shorter overall survival. We therefore concluded that increased densities of ST2-postive cells relate to Treg accumulation within the adenoma/CRC microenvironment, suggesting the IL-33/ST2 pathway as a potential contributor for immunosuppressive milieu formation that impact disease stage and prognosis in patients with CRC.

Tumoral Expression of CD166 in Human Esophageal Squamous Cell Carcinoma: Implications for Cancer Progression and Prognosis.

Accumulating data showed that cancer stem cells (CSCs) identified by cell surface markers contribute to the initiation, progression, and prognosis of human cancers. In this study, the expression of CSC candidates CD166, CD44, and Lgr5 in 65 cases of esophageal squamous cell carcinoma (ESCC) and 16 cases of control esophageal tissues were examined with immunohistochemistry (IHC). The correlation between tumoral expression levels of these CSC candidates and clinicopathological variables was analyzed. IHC results showed that the expression of CD166 in esophageal control tissues was completely negative, but it was in 87.69% (57/65) ESCC tissues. The expression of CD44 and Lgr5 did not differ between esophageal control tissues and ESCC tissues (p > 0.05). In addition, there were not correlations found among the expression levels of CD166, CD44, and Lgr5 in ESCC tissues. Clinicopathological analysis revealed that the tumoral expression level of CD166 correlated with lymph node involvement and TNM staging in patients with ESCC, and lower tumoral expression of CD44 was found in patients with advanced TNM staging. Kaplan-Meier survival curves suggested that expression level of CD166 appeared to have a negative impact on overall survival rate after surgery in patients with ESCC. Such impact was not found in other two CSC candidates. The authors therefore conclude that CD166 is a potential prognostic biomarker and correlates with advanced progression features in patients with ESCC.

IL-33 in the tumor microenvironment is associated with the accumulation of FoxP3-positive regulatory T cells in human esophageal carcinomas.

Forkhead box p3 (Foxp3+) regulatory T cells (Tregs) are abundant in the tumor microenvironment where they dampen functions of host anti-tumor immunity and promote cancer progression. Cytokine signaling is essential for the generation and function maintenance of Tregs in patients with cancers. Recent in vitro and in vivo studies have described that interleukin (IL)-33 plays a critical role in regulating the expansion and function of Tregs. However, the regulatory role of IL-33 in Treg recruitment within the microenvironment of human esophageal squamous cell carcinoma (ESCC) to date is poorly understood. In this study, we have therefore characterized the expression of IL-33 by immunohistochemistry (IHC) and double immunofluorescence staining and analyzed its relationship with FoxP3+ Treg accumulation in the microenvironment in 80 patients with ESCC. IHC observation revealed a high expression level of IL-33 in both ESCC mass and stroma, which paralleled to a high density of FoxP3+ Tregs accumulated in the same compartments. Statistical analysis showed that the scores for cell densities of tumor- and stroma-expressing IL-33 were significantly correlated with the scores for density of FoxP3+ Tregs in the tumor stroma. Further immunofluorescence images demonstrated that IL-33 functional receptor, ST2, was preferentially expressed in FoxP3+ Tregs, suggesting a possible effecting pathway for IL-33. In addition, cyclooxygenase-2, one of the important immunosuppressive factors, was highly illustrated in FoxP3+ Tregs. We have therefore concluded that microenvironmental-expressing IL-33 is associated with the recruitment of Tregs in human ESCCs.

Contribution of IL-33 to the Pathogenesis of Colorectal Cancer.

The development of colorectal cancer (CRC) is not only determined by transformed cells per se, but also by factors existing in their immune microenvironment. Accumulating scientific evidence has revealed that interleukin (IL)-33, an IL-1 family member, plays an essential role in the regulation of immune response and is relevant in CRC pathogenesis. Data from both human and experimental studies demonstrated that IL-33 inhibits host anti-tumor immunity, remodels tumor stroma and enhances angiogenesis, thereby promoting the development of CRC. These pro-tumor effects of IL-33 are mainly mediated by IL-33 receptor ST2 (also known as IL-1RL1). Based on those findings, it is currently hypothesized that the IL-33/ST2 pathway is a potential biomarker and therapeutic target for colorectal tumorigenesis. Herein, we summarize the recent discoveries in understanding the critical role of the IL-33/ST2 pathway in contributing to the pathogenesis of colorectal tumorigenesis and discuss its potential implications for the future development of effective anti-tumor strategies.

Cellular and clinicopathological features of the IL-33/ST2 axis in human esophageal squamous cell carcinomas.

BACKGROUND: Emerging evidence has suggested that interleukin (IL)-33 and its primary functional receptor ST2 are involved in the pathogenesis of tumorigenesis. METHODS: Using immunohistochemistry (IHC) and double immunofluorescence staining, we characterized the cellular and clinicopathological features of the IL-33/ST2 axis in different compartments in human esophageal squamous cell carcinoma (ESCC) surgical specimens. RESULTS: IHC data revealed an increased expression of IL-33-immunoreactivity (IR) and ST2-IR located in both ESCC cells and tumor stromal cells; which were associated with advanced clinicopathological features such as TNM stages and node involvement. However, the Kaplan-Meier analysis showed that densities of neither IL-33 positive nor ST2 positive cells in both the ESCC mass and stroma were associated with the overall survival rate in patients with ESCC. Double immunofluorescence staining for cellular feature analysis demonstrated that these IL-33 positive and ST2 positive cells in ESCCs were with a high proliferation rate, and IL-33-IR was frequently co-expressed with ST2-IR in both ESCC and stromal cells. CONCLUSION: Significant altered cellular features of the IL-33/ST2 axis in ESCCs were associated with advanced clinicopathological variables. The data suggest that the IL-33/ST2 axis might be involved in the progression of human ESCCs.

A Systematic Review of Epidemiology and Risk Factors Associated With Chinese Inflammatory Bowel Disease.

Background: Recent epidemiological data have revealed a dramatically rising prevalence and incidence of inflammatory bowel diseases (IBDs) in Mainland China, a rapidly growing industrialized region, over the last two decades. Objectives: We performed a systematic review to investigate the changing trends in the incidence of IBD in Mainland China and summarized the recent findings in risk factors associated with Chinese IBD. Methods: Relevant references were obtained from an electronic database search via MEDLINE and EMBASE (for English literatures), the China Academic Journals Full-text Database (CJFD) and the China Science Periodical Database (CSPD) in Wanfang Data (for Chinese literatures). Results: Total 1,584 abstracts in Chinese and 171 abstracts in English were collected. Eight full-text with epidemiological data, 25 with risk factor data in Chinese and 7 full-text with epidemiological data, 12 with risk factor data in English were finally identified and included for analysis. Data from included epidemiological studies has showed a striking increase in the incidence rate of IBD in Mainland China over time, and current incidence rates for IBD, UC, and CD were 1.80 (IBD), 1.33 (UC), and 0.46/1,000,000 (CD), though it varies among regions and ethnic minority populations. In addition, several risk factors including environmental factors, diet, intestinal infectious agents, hygiene, stress, and lifestyle have been reported to be associated with the increased incidence of Chinese IBD. Conclusion: This systematic review revealed an increased incidence of IBD in Mainland China. Although it is still lower than that in the Western world, however, China has a huge population; therefore, the total number of IBD patients might not be so little as previously thought and the disease burden of IBD in China is likely underestimated. HIGHLIGHTS Recent epidemiological data have revealed a dramatically rising prevalence and incidence of inflammatory bowel diseases (IBD) in Mainland China, a rapidly growing industrialized region, over the last two decades.This systematic review based on recent epidemiological data has revealed a striking increase in the incidence rate of IBD in Mainland China, though it varies among regions and ethnic minority populations.Several potential risk factors of IBD including environmental factors, diet, intestinal infectious agents, hygiene, stress, and lifestyle have been reported to be associated with the increased incidence of Chinese IBD.This systematic review on epidemiologic and risk factor studies has expanded understanding of the occurrence, distribution, geographic variance and risk factors of Chinese IBD and will provide clinicians important information in understanding current status of IBD in Mainland China.

IL-1ß/IL-6 network in the tumor microenvironment of human colorectal cancer.

OBJECTIVES: Recent studies suggest that the interaction between interleukin (IL)-1ß and IL-6 in the microenvironment might be involved in the development and progression of human colorectal cancer (CRC). However, the expression of IL-1ß/IL-6 network within the CRC microenvironment is not fully understood. MATERIALS AND METHODS: The level of IL-1ß/IL-6 network expression in 40 biopsies of sporadic CRC and 15 biopsies of controls was assessed using quantitative real-time polymerase chain reaction (PCR) assay, immunohistochemistry (IHC) and double immunofluorescence staining. RESULTS: Quantitative results obtained by real-time PCR revealed that both IL-1ß and IL-6 mRNA expressions were increased in CRC tissues compared with expressions in controls. In which, IL-6 mRNA expression in primary CRC tissues showed a statistically significant relationship with tumor invasion depth. IHC observations confirmed that increased expression of IL-1ß and IL-6 immunoreactivities was located in both the CRC epithelium and stroma. Furthermore, IHC results also revealed that increased expression of IL-1ß receptor type 1 (IL-1R1) and IL-6 receptor (IL-6R) were observed in both CRC epithelial and stromal cells. IHCs in serial CRC sections and double immunofluorescence staining revealed a highly co-expression of IL-1R1 immunoreactivity with IL-6 immunoreactivity in the same cells, which confirmed a histological fundament of IL-1ß/IL-6 network. CONCLUSION: The IL-1ß/IL-6 network is highly expressed in the CRC microenvironment, indicating that this network is important in the progression of CRC.

Tumor-Associated Fibroblasts and Microvessels Contribute to the Expression of Immunosuppressive Factor Indoleamine 2, 3-Dioxygenase in Human Esophageal Cancers.

Recent studies have provided considerable evidence to support the hypothesis that tumor stroma plays a crucial role in the induction of immune tolerance to human cancers. Here, we investigated the contribution of reactive stromal tumor-associated fibroblasts (TAFs) and microvessels to the immunosuppressive factor indoleamine 2,3-dioxygenase (IDO) expression in the ESCC microenvironment. The immunohistochemical (IHC) analyses demonstrated a significant increased densities of TAFs and microvessels in the ESCC stroma, double IHCs showed that these increased TAFs and microvessels were with a high proliferation activity. Further IHC examinations revealed that increased expression of IDO were frequently observed in the stromal cells with TAF morphology and microvessels. Double immunofluorescence examinations confirmed the colocalization of IDO positive cells with SMA-alpha positive TAFs and CD34 positive endothelial cells in the ESCC stroma. Our current findings strongly suggest that the activated stromal TAFs and endothelial cells of microvessels contribute to the expression of IDO and then the orchestration of immunosuppressive microenvironment.

Increased expression of interleukin-21 along colorectal adenoma-carcinoma sequence and its predicating significance in patients with sporadic colorectal cancer.

The role and significance of interleukin (IL)-21 in the development of sporadic CRC have not been well defined. The aim of this study is therefore to investigate the dynamics of the IL-21 along colorectal adenoma-carcinoma sequence and to evaluate the impact of IL-21 on clinicopathological parameters and CRC prognosis. The real-time PCR results showed that the level of IL-21 in adenomas (n=50) and sporadic CRC (n=50) were significantly higher than that in normal controls (n=18), which were predominately observed in the adenoma/CRC stroma. Analysis revealed that IL-21 level was correlated with the overall survival time in CRC patients. Double immunofluorescence observations confirmed that IL-21 positive cells were mostly natural killer cells and T lymphocytes in the tumor stroma. These results indicate that significant increased IL-21 expression present within the adenoma/CRC microenvironment might have a potential predicating significance for survival time in patients with CRC.

Temporal and spatial changes of cells positive for stem-like markers in different compartments and stages of human colorectal adenoma-carcinoma sequence.

Considerable evidence supports the idea that stem-like cells may play an essential role during the development of colorectal cancer (CRC). To accomplish this aim, we use immunohistochemistry (IHC) and double IHC with different potential stem-like markers, anti-musashi (Msi), anti-CD133, anti- LGR5 and anti-ALDH1 to examine the presentation of stem-like cells in different compartments including adenoma/CRC epithelium, transitional crypts and tumor stroma in colorectal adenoma and CRC. The results showed that cells positive for stem-like markers were remarkably increased in number and frequently observed in the adenoma/CRC epithelium, transitional crypts and tumor stroma. Notably, the population of cells positive for stem-liker markers was expanded from the base to the middle part of the transitional crypt in both adenoma and CRC tissues, reflecting that stem-like cells are likely involved in the process of colorectal tumorigenesis. Counting results showed that the grading scores of cells positive for LGR5 and ALDH1 in the adenoma/CRC epithelium were significantly increased relative with the control epithelium, and associated with the degree of dysplasia in the adenoma and node involvement in the CRC (all P < 0.05). In addition, the density of cells positive for stem-like markers in the adenomatous/cancerous stroma was also increased and paralleled an increase in the density of proliferative stromal cells labeled by PCNA, which were primarily identified as vimentin positive fibroblasts. Our results have revealed a changed temporal and spatial presentation of stem-like markers in different stages of human colorectal adenoma-carcinoma sequence, which might be a hallmark of the adenoma-carcinoma transition.

Accumulation of FoxP3+ T regulatory cells in the tumor microenvironment of human colorectal adenomas.

OBJECTIVES: T regulatory cells (Tregs) play a critical important role for the occurrence and development of human tumors. Most human colorectal cancers (CRCs) develop from the preformed adenomas, this study is therefore designed to evaluate forkhead box P3 (FoxP3)+ Tregs in human colorectal adenomas. MATERIALS AND METHODS: FoxP3+ Tregs in human colorectal adenomas were evaluated with immunohistochemistry (IHC) and real-time PCR, and compared to CRCs and normal tissues. In addition, the change of Treg immunosuppressive cytokine interleukin (IL)-10 was examined with IHC and real-time PCR. RESULTS: increased FoxP3+Tregs were observed in the adenomatous stroma/epithelium and the density in colorectal adenomas, which was similar to that in the CRCs, significantly increased as compared to normal tissues. Numerous IL-10+cells were observed in the adenomatous stroma, but not in adenomatous epithelium, as compared with the controls. The density grading score of IL-10+ cells in the adenomas confirmed an increased density of IL-10+cells in the adenomatous/CRC stroma. Double IHCs with CD4/CD25 and IL-10/FoxP3 antibodies confirmed above observations and revealed that IL-10 was at least partially released from increased Tregs. Quantitative real-time PCR results confirmed that the expression levels of FoxP3 and IL-10 mRNAs in the adenomas were increased, which equivalent to that in the CRCs. CONCLUSION: accumulation of FoxP3+ Tregs in the tumor microenvironment is an early event along the adenoma-carcinoma sequence, and might play a role in the regulation of host immune response to the initiation of CRC.

IL-9 antibody injection suppresses the inflammation in colitis mice.

Diverse T help (Th) cells play a crucial role in the processing and maintaining of chronic inflammation as seen in ulcerative colitis (UC). Th9, a novel subset of Th cells that primarily produces interleukin (IL)-9, has recently been associated with the development of inflammatory diseases. In this study, we evaluated the presentation of Th9 cells in inflamed tissues of human and experimental mouse UC, and examined the therapeutic efficiency of anti Th9 cytokine IL-9 in the experimental mouse UC. Using immunohistochemistry (IHC), we evaluated the presentation of Th9 cells labelled by transcriptional factor PU.1 in both human and dextran sulfate sodium (DSS) induced mouse colitis biopsies. The results showed that increased PU.1 positive Th9 cells were mainly located in the lamina propria in relative with the controls, intraepithelial Th9 cells can also be observed but at low density. Double IHCs revealed that most of PU.1 positive cells were CD3 positive lymphocytes in human UC specimens. Anti-IL-9 antibody injection for 2 weeks reduced the severity of inflammation in DSS induced colitis mice. Our results suggest that The Th9/IL-9 is involved in the pathogenesis of UC.

Elevated proinflammatory cytokine IL-17A in the adjacent tissues along the adenoma-carcinoma sequence.

Considerable evidence has suggested that chronic inflammation is a causative factor in the development of human colorectal cancer (CRC). Interleukin (IL)-17A produced mainly by Th17 cells is a novel proinflammatory cytokine and increased IL-17A is associated with colorectal neoplastic transformation. In this study, we have evaluated the expression of IL-17A in the adjacent tissues along the colorectal adenoma-carcinoma sequence. The expression of IL-17A in the adjacent tissues of colorectal adenoma (adenoma-adjacent, n = 32) and sporadic CRC (CRC-adjacent, n = 45) was examined. In addition, the expression pattern of Th17 cell differentiation stimulators (IL-1ß, IL-6 and IL-23A) in the adjacent tissues were also examined. The results showed that the expression level of IL-17A mRNA was non-statistically increased (4-fold higher) in the adenoma-adjacent tissues and it became significantly increased (9-fold higher) in the CRC-adjacent tissues as compared with the control. The expression level of IL-17A in the CRC-adjacent tissues was not associated with CRC clinicopathological parameters and overall survival. Immunohistochemistry confirmed an increased density of intraepithelial IL-17A expressing cells in the CRC-adjacent tissues. The Th17 cell differentiation simulators IL-1ß and IL-6 were also shown in an increase trend from the adenoma-adjacent to CRC-adjacent tissues. These results provide evidence that IL-17A/Th17 response is enhanced in the adjacent tissues during the colorectal neoplastic transformation.

IL-17A in the tumor microenvironment of the human colorectal adenoma-carcinoma sequence.

OBJECTIVES: Interleukin (IL)-17A is an important pro-inflammatory cytokine and involved in the colorectal carcinogenesis. In this study, the authors evaluated the dynamic change of IL-17A expression in the tumor microenvironment throughout the colorectal adenoma-carcinoma sequence. MATERIALS & METHODS: Using quantitative real-time PCR (polymerase chain reaction) and semi-quantitative immunohistochemistry, the authors examined the expression level of IL-17A in 50 of human colorectal adenoma tissues, 50 of colorectal cancer (CRC) tissues and 15 controls. The relationship between IL-17A expression and clinicopathological parameters throughout the sequence was also evaluated. RESULTS: The results revealed a step-up increased IL-17A mRNA level throughout the colorectal adenoma-carcinoma sequence, which began to increase in the adenomas and became even higher in the CRCs; notably, the increase of IL-17A mRNA level in the adenomatous tissues was associated with the severity of dysplasia. Immunohistochemical analysis confirmed the real-time PCR results and revealed gradually increasing IL-17A cells in both the stroma and adenomatous/cancerous epithelium. In addition, the quantitative real-time PCR result has also revealed an increased expression of TH17-stimulating factors throughout the sequence. CONCLUSIONS: IL-17A and TH17 are highly activated throughout the colorectal adenoma-carcinoma sequence.

Progressive cellular response in the lamina propria of the colorectal adenoma-carcinoma sequence.

AIMS: The lamina propria is inevitably involved in epithelial transformation. The aim was to evaluate the dynamic cellular changes in the tumour lamina propria throughout the colorectal adenoma-carcinoma sequence. METHODS AND RESULTS: Using immunohistochemistry and double immunohistochemistry, we examined lamina propria cellular changes in 41 colorectal adenomas, 25 colorectal cancers and 15 control tissues. The results showed that the proliferation labelling index in lamina propria cells began to increase in the precancerous lesions (adenomas) and became even higher in the colorectal cancers; these proliferative cells were primarily identified as myofibroblasts and lymphocytes. Phenotypic analysis revealed gradually increasing lymphocytic infiltration in both the lamina propria and adenomatous epithelium, as well as myofibroblasts in the lamina propria. However, the intraepithelial macrophage density also showed a tendency to increase gradually. Furthermore, cyclooxygenase-2-expressing cell density and microvessel density gradually increased in the tumour lamina propria throughout the adenoma-carcinoma sequence. CONCLUSIONS: Progressive cellular responses in the lamina propria could be involved in the adenoma-carcinoma transition.

Dynamic changes of interleukin-8 network along the colorectal adenoma-carcinoma sequence.

The interleukin-8 (IL-8) network is involved in the colorectal cancer (CRC) progression. However, its role during the adenoma-carcinoma transition to date has not been fully investigated. To evaluate the dynamic changes of IL-8 network along the colorectal adenoma-carcinoma sequence, we examined the tissue IL-8 mRNA level in colorectal biopsies from 53 colorectal adenomas, 44 CRCs and 18 controls by quantitative real-time PCR (Q-PCR), and the expressions of IL-8 and its receptors (IL-8RA and IL-8RB) in the tumor microenvironment by immunohistochemistry (IHC) and double IHCs. The results showed that the tissue IL-8 mRNA level began to increase in the precancerous lesions (adenomas) as compared with the controls and became even higher in the CRCs. Significantly, the increase of IL-8 mRNA levels was associated with the increase of dysplastic grades in the adenomas, and also paralleled to the increase of Duke's stages in the CRCs. IHC results revealed that IL-8 and its receptors, IL-8RA and IL-8RB, were observed both in the stroma and in the adenomatous/cancerous cells. By double IHCs, the IL-8 expression was characterized in macrophages, lymphocytes and myofibroblasts in the tumor stroma. Further double IHC identified the co-expression of IL-8 receptors (IL-8RA and IL-8RB) with CD34 positive tumor-associated microvessels in both the adenomas and CRCs. We, therefore, conclude that activated IL-8 network in the tumor microenvironment may function as a significant regulatory factor for the adenoma progression and the adenoma-carcinoma transition.

Dendritic cell infiltration pattern along the colorectal adenoma-carcinoma sequence.

We have previously reported that the dendritic cell (DC) functional index cytokine interleukin-12 was significantly decreased in colorectal carcinoma (CRC) tissues. In this study, the DC infiltration pattern and the density of mature DCs (mDCs; labeled by anti-CD83 and anti-CD208) and immature DCs (iDCs; labeled by anti-CD1alpha) were characterized using immunohistochemistry (IHC) in tissue samples from 23 patients with CRC, 33 patients with colorectal adenoma (CRA), and 19 healthy controls. In addition, the DC function inhibitor cyclooxygenase-2 (COX-2) and the downstream signal molecule prostaglandin E2 (PGE2) and related receptors EP2/EP4 were measured by quantitative real-time PCR and double immunofluorescence staining. The IHC analyses revealed changed densities of mDCs and iDCs in the tumor microenvironment; in CRA and CRC, the density of mDCs was decreased, but the density of iDCs was gradually increased. Furthermore, the distribution patterns of DCs were also altered. In CRA, mDCs were abundantly distributed in the subepithelial stroma of the adenomatous mass. In CRC, the distribution of mDCs in the tumor stroma was not homogeneous, and mDCs residing in the stroma at invading edges were more frequently found compared with in the intratumoral stroma (P<0.05). Increased iDCs were found in the intratumoral mass in CRC, and some infiltrated into the malignant epithelium. By quantitative real-time PCR, a gradually increased level of COX-2 mRNA was demonstrated in the local tissues along the adenoma-carcinoma sequence, and double immunofluorescence staining showed a colocalization of PGE2 receptors EP2/EP4 with mDCs in the stroma of CRC. In conclusion, our current findings revealed an altered DC infiltration pattern along the adenoma-carcinoma sequence; gradually increased COX-2 expression might contribute to the DC functional defect.

Immunohistochemical examination of gastrin, gastrin precursors, and gastrin/CCK-2 receptor in human esophageal squamous cell carcinomas.

A promoting effect of gastrin on stimulating Barrett's oesophagus proliferation has been demonstrated, but whether it plays a regulating role for esophageal squamous cell carcinoma (ESCC) to date has not been fully investigated. The aim of this study is to examine the expressions of gastrin, gastrin precursors and gastrin/CCK-2 receptor in ESCC. Tissue specimen sections from 38 patients with ESSC obtained from a high incidence area of north China were assessed using immunohistochemistry for amidated gastrin, gastrin precursors (progastrin and glycine-extended gastrin) and gastrin/CCK-2 receptors. Their clinical histopathological significance was also analyzed. Of 38 ESCC, the immunoreactivities of gastrin, glycine-extended gastrin and progastrin were observed in 13.2% (5/38), 7.9% (3/38) and 23.68% (9/38) cases. The expression of progastrin was obviously higher than other gastrins, though not significantly (P > 0.05). In positive cases for gastrin or glycine-extended gastrin, the scores of positive tumor cell numbers were at a lower density (<10/abundant-distributed field). However, the scores of progastrin positive tumor cell density in five of nine positive cases were over 10/abundant-distributed field. The immunoreactivity of gastrin/CCK-2 receptor was also observed in 15.8% (6/38) ESCC cases. There was not significant correlation regarding immunohistochemical results with known histomorphological parameters i.e. gender, tumor location and TNM stages. Based on our current results, ESCC tumor cells could be a possible cellular source of gastrin precursors, which has been postulated to play a role in regulating the growth in some human tumor cells.