Intranodal Mapping Using Carbon Dye Results in More Accurate Lymph Node Staging in Colon Cancer Patients.

INTRODUCTION: Small nodal tumor infiltrates (SNTI)-defined as isolated tumor cells and micrometastases-are associated with worse disease-free and overall survival in stage I and II colon cancer patients. Their detection, however, remains challenging. The objective of the present study was to evaluate whether there is a correlation between the location of SNTI and phagocytosed carbon dye particles in sentinel lymph nodes (SLN) of colon cancer patients. MATERIALS AND METHODS: Isosulfan blue and carbon dye were injected intraoperatively near the tumor to mark the SLN. Serial sections of SLN were stained with hematoxylin-eosin and immunohistochemistry. Intranodal distribution of phagocytosed carbon particles was compared to the presence of SNTI. RESULTS: Of a cohort of 159 patients, 24 patients had SNTI in their lymph nodes (LN). SNTI were found in a total of 116 LN of which 66 were SLN and 50 were non-SLN. In 59, these 116 LN with SNTI phagocytosed carbon dye were found (50.9 %). Phagocytosed carbon dye was identified significantly more often in SLN (49 of 66 SNTI positive SLN) compared to 10 of 50 SNTI positive non-SLN (p < 0.001). In 52 out of 59 LN (88.1 %), phagocytosed carbon dye was in close proximity to SNTI. CONCLUSIONS: In the majority of patients, SNTI are located in the same SLN compartment as phagocytosed carbon dye particles. Our investigation provides evidence that the use of carbon dye facilitates SNTI detection and improves LN staging in colon cancer. Therefore, the concept of intranodal mapping-which has been previously described for melanoma-can be extended to colon cancer patients.

Blue dye injection does not induce dissemination of epithelial cells during SLN procedure in colon cancer patients.

INTRODUCTION: The sentinel lymph node (SLN) procedure for colon cancer patients has been increasingly performed over the past decade and has shown advantages regarding lymph node staging. However, there are concerns that the manipulation of the colon, particularly the blue dye injection, results in isolated tumor cell dissemination to lymph nodes. Therefore, the objective of the present study was to evaluate whether the blue dye injection during the SLN procedure for colon cancer induces epithelial cell dissemination to the regional lymph nodes using a fake SLN procedure as a model. METHODS: One hundred seventy-four colon cancer patients underwent open oncologic colon resection and SLN procedure according to a standardized protocol. For the fake SLN procedure, blue dye was injected ex vivo, into the subserosa of a nontumor-bearing segment of the resected colon in 37 unselected patients. Three levels of each SLN were stained with H&E and with the pancytokeratin marker AE1/AE3 and were analyzed for the presence of cytokeratin positive cells. RESULTS: Identification of fake SLN was successful in 32 of the 37 patients (86 %). Seventy fake SLN were histologically confirmed. The median number of fake SLN was 2 per patient (range 1-8). None of the fake SLN showed any disseminated epithelial cells. CONCLUSIONS: The present prospective study provides compelling evidence that blue dye injection during sentinel lymph node procedure for colon cancer does not induce epithelial cell dissemination to the sentinel lymph nodes. Therefore, isolated tumor cells in sentinel lymph nodes result from a true metastatic process.

Sentinel lymph node procedure leads to upstaging of patients with resectable colon cancer: results of the Swiss prospective, multicenter study sentinel lymph node procedure in colon cancer.

BACKGROUND: The value of the sentinel lymph node (SLN) procedure in colon cancer patients remains a matter of debate. The objective of this prospective, multicenter trial was 3-fold: to determine the identification rate and accuracy of the SLN procedure in patients with resectable colon cancer; to evaluate the learning curve of the SLN procedure; and to assess the extent of upstaging due to the SLN procedure. METHODS: One hundred seventy-four consecutive colon cancer patients were enrolled onto this prospective trial. They underwent an intraoperative SLN procedure with isosulfan blue 1% injected peritumorally followed by open standard colon resection with oncologic lymphadenectomy. Three levels of each SLN were stained with hematoxylin and eosin (H&E) and immunostained with the pancytokeratin marker AE1/AE3 if H&E was negative. RESULTS: SLN identification rate and accuracy were 89.1% and 83.9%, respectively. SLN were significantly more likely to contain tumor infiltrates than non-SLN (P < 0.001). Both SLN identification rate (P = 0.021) and the sensitivity of the procedure (P = 0.043) significantly improved with experience. The use of immunohistochemistry in SLN resulted in an upstaging of 15.4% (16 of 104) stage I and II patients considered node-negative in initial H&E analysis. CONCLUSIONS: The SLN procedure for colon cancer has good identification and accuracy rates, which further improve with increasing experience. Most importantly, the SLN procedure results in upstaging of >15% of node-negative patients. The potential advantage of performing the SLN procedure appears to be particularly important in these patients because they may potentially benefit from adjuvant therapy.

Abdominal actinomycosis: a rare complication after laparoscopic gastric bypass.

A 33-year-old, morbidity obese woman underwent a laparoscopic Roux-en-Y gastric bypass in November 2004. She presented 18 months later with a history of recurrent pain in the upper region of the abdomen and severe vomiting. Radiologic and endoscopic evaluations revealed wall thickening in the transverse colon and a solid tumor near the liver. Therefore, a sonography-guided biopsy of the tumor was performed. Cytopathological examination revealed actinomycosis. Thus, therapy with penicillin was started, after which the parameters associated with the infection decreased. The symptoms persisted, however, and the decision was made to operate on the patient to resect the abdominal masses. Nearly 90% of the masses could be removed. Histological analysis showed a fibro-productive inflammation with an actinomycotic etiology. Antibiotic therapy with penicillin was continued for 6 months. Actinomycosis must be considered in the differential diagnosis of patients with abdominal mass, wall thickening of the intestine, and other such symptoms, including abdominal pain following bariatric surgery, even many years after the intervention.

Carbon dye staining of sentinel lymph nodes facilitates microstaging of colon cancer patients.

BACKGROUND: Carbon dye, when peritumourally injected, permanently marks the drainage site of sentinel lymph nodes (SLN). The objective of the current study was to evaluate whether the use of carbon dye facilitated the detection of small nodal tumour infiltrates in colon cancer patients. METHODS: In a prospective trial, 19 patients underwent open, oncological resections of localized colon cancer and SLN procedure according to a standardized protocol. Isosulfan blue 1% and sterile filtered carbon dye (mixed 1:1) were injected into the subserosa circumferentially around the tumour. Lymph nodes staining blue were marked as SLN. Serial sections of each SLN were stained with hematoxylin and eosin (H&E) and with the pancytokeratin marker AE1/AE3. The intranodal presence and site of carbon particles were noted and compared with the location of possible tumour infiltrates. RESULTS: Identification of at least one SLN was successful in 18 patients (identification rate 95%). Four patients (22%) were pN+, 11 (61%) were pN0(i-). Three patients (17%) were upstaged from pN0(i-) to pN0(i+) as isolated tumour cells were detected in their SLN: in two (11%) of the three patients, carbon dye and isolated tumour cells were found in the same nodal compartment, hence facilitating the recognition of isolated tumour cells by the pathologist. CONCLUSION: The use of carbon dye in the SLN procedure for colon cancer may facilitate the detection of small nodal tumour infiltrates.

Kupffer cell-expressed membrane-bound TNF mediates melphalan hepatotoxicity via activation of both TNF receptors.

Isolated hepatic perfusion of nonresectable liver cancer using the combination of TNF and melphalan can be associated with a treatment-related hepatotoxicity. We investigated whether, apart from TNF, also melphalan is cytotoxic in primary murine liver cells in vitro and investigated mediators, mode of cell death, and cell types involved. Melphalan induced a caspase-dependent apoptosis in hepatocytes, which was not seen in liver cell preparations depleted of Kupffer cells. Neutralization of TNF prevented melphalan-induced apoptosis and liver cells derived from mice genetically deficient in either TNFR 1 or 2, but not from lpr mice lacking a functional CD95 receptor, were completely resistant. Cell-cell contact between hepatocytes and Kupffer cells was required for apoptosis to occur. Melphalan increased membrane-bound but not secreted TNF in Kupffer cells and inhibited recombinant TNF-alpha converting enzyme in vitro. Melphalan induced also severe hepatotoxicity in the isolated recirculating perfused mouse liver from wild-type mice but not from TNFR 1 or 2 knockout mice. In conclusion, this study shows that melphalan elicits membrane TNF on Kupffer cells due to inhibition of TNF processing and thereby initiates apoptosis of hepatocytes via obligatory activation of both TNFRs. The identification of this novel mechanism allows a causal understanding of melphalan-induced hepatotoxicity.

Immunohistochemical analysis reveals high frequency of PMS2 defects in colorectal cancer.

BACKGROUND & AIMS: Germline mutations in the DNA mismatch repair (MMR) genes MSH2, MSH6, or MLH1 predispose to colorectal cancer (CRC) with an autosomal dominant inheritance pattern. The protein encoded by PMS2 is also essential for MMR; however, alterations in this gene have been documented only in extremely rare cases. We addressed this unexpected finding by analyzing a large series of CRCs. METHODS: Expression of MSH2, MSH6, MLH1, and PMS2 was studied by immunohistochemistry in 1048 unselected, consecutive CRCs. Where absence of MMR proteins was detected, microsatellite instability and cytosine methylation of the respective gene promoter were analyzed. The DNA of patients presenting with PMS2-deficient cancers was examined for germline and somatic alterations in the PMS2 gene. RESULTS: An aberrant pattern of MMR protein expression was detected in 13.2% of CRCs. Loss of expression of MSH2, MSH6, or MLH1 was found in 1.4%, 0.5%, and 9.8%, respectively. PMS2 deficiency accompanied by microsatellite instability was found in 16 cases (1.5%) with a weak family history of cancer. The PMS2 promoter was not hypermethylated in these cases. Despite interference of the PMS2 pseudogenes, we identified several heterozygous germline mutations in the PMS2 gene. CONCLUSIONS: PMS2 defects account for a small but significant proportion of CRCs and for a substantial fraction of tumors with microsatellite instability. However, the penetrance of heterozygous germline mutations in PMS2 is considerably lower than that of mutations in other MMR genes. The possible underlying causes of this unorthodox inheritance pattern are discussed.

Alterations of RB1, p53 and Wnt pathways in hepatocellular carcinomas associated with hepatitis C, hepatitis B and alcoholic liver cirrhosis.

Major etiologic factors associated with human hepatocellular carcinomas (HCCs) include infection with hepatitis C (HCV) and hepatitis B virus (HBV), excess alcohol intake and aflatoxin B(1) exposure. While the G-->T p53 mutation at codon 249 has been identified as a genetic hallmark of HCC caused by aflatoxin B(1), the genetic profile associated with other etiologic factors appears to be less distinctive. In our study, we screened HCCs resulting from HCV infection (51 cases), HBV infection (26 cases) or excess alcohol intake (23 cases) for alterations in genes involved in the RB1 pathway (p16(INK4a), p15(INK4b), RB1, CDK4 and cyclin D1), the p53 pathway (p53, p14(ARF) and MDM2) and the Wnt pathway (beta-catenin, APC). Alterations of the RB1 pathway, mainly p16(INK4a) methylation, loss of RB1 expression and cyclin D1 amplification, were most common (69-100% of cases). There was a significant correlation between loss of RB1 expression and RB1 methylation. All 24 HCCs with RB1 promoter methylation lacked RB1 expression, while none of the 67 cases with RB1 expression exhibited RB1 methylation (p < 0.0001), suggesting that promoter methylation is a major mechanism of loss of RB1 expression in HCCs. Alterations of the p53 pathway consisted mostly of p53 mutations or p14(ARF) promoter methylation (20-48%). Mutations of the p53 gene were found at a similar frequency (13-15%) in all etiologic groups, without any consistent base change or hot spot. Mutations of beta-catenin were found in 13-31% of cases, while no APC mutations were detected in any of the HCCs analyzed. With the exception of only 3 of 39 cases (8%), cyclin D1 amplification and beta-catenin mutations were mutually exclusive, supporting the view that cyclin D1 is a target of the Wnt signaling pathway. Overall, the RB1, p53 and Wnt pathways were commonly affected in HCCs of different etiology, probably reflecting common pathogenetic mechanisms, i.e., chronic liver injury and cirrhosis, but tumors associated with alcoholism had more frequent alterations in the RB1 and p53 pathways than those caused by HCV infection.

Evaluation of hepatitis B and hepatitis C virus-infected renal allograft recipients with liver biopsy and noninvasive parameters.

BACKGROUND: Patients with end-stage renal disease are at high risk for hepatitis B (HBV) or hepatitis C virus (HCV) infection. Because therapy indication for viral hepatitis depends on virologic, biochemical, and histologic criteria, liver biopsy usually is necessary. Recently, a panel of serum fibrosis markers has been postulated to allow quantification of liver fibrosis by noninvasive means. METHODS: A cross-sectional study of all hepatitis B surface antigen (HBsAg)- and anti-HCV-positive renal allograft recipients among 900 renal allograft recipients regularly controlled in the authors' outpatient nephrology service was performed. The correlation between histologic, biochemical, and virologic parameters was assessed with an emphasis on the fibrosis marker hyaluronate in this immunosuppressed population. RESULTS: Twenty-two HBsAg- and 62 anti-HCV-positive patients were analyzed. Based on polymerase chain reaction results, 86% of anti-HCV-positive and 95% of HBsAg-positive patients had actively replicating infection. In 41 of 67 (61%) patients with replicating disease, liver biopsy was performed, and the association of various biochemical parameters with the histologic scores for necroinflammation and fibrosis was investigated. Less than 10% of these patients had advanced fibrosis, although the mean time of infection was more than 15 years. We found no correlation of any of the serum parameters (including hyaluronate) with histologic activity of liver disease except for the peak glutamate-oxalacetate transaminase value recorded during the entire posttransplant period. CONCLUSION: Liver biopsy remains the gold standard for evaluation of liver disease and therapy decision in immunosuppressed renal allograft recipients.