Alessandro Balani

Effects of formalin, methacarn, and fineFIX fixatives on RNA preservation.

Formalin-fixed tissues represent the most abundant clinical material for retrospective studies. However, formalin highly affects macromolecules, impairing their extraction and analysis. In this study, the suitability of some potential substitutes of formalin for RNA-based applications has been considered. Conventional formalin was compared with methacarn and the commercial FineFIX. Their impact on overall RNA preservation was investigated in a cell line-based model fixed during a time course treatment and in a series of fixed human tissues. RNA yield was detected by Nanodrop; ribosomal RNA (rRNA) integrity by electrophoresis and the Agilent Bioanalyzer; messenger RNA (mRNA) integrity by Northern blot and endpoint reverse transcription-polymerase chain reaction; and mRNA amount by real-time polymerase chain reaction. In the cell line model, formalin fixation showed time-dependent detrimental effects on overall RNA preservation. Methacarn and FineFIX were more conservative on both rRNA and mRNA preservation and their impact was time-independent. In tissues, high rRNA degradation levels were found in all fixed specimens, contrasting with the results found in the cells. Conversely, the effects of the fixatives on mRNA integrity reflected the observations shown in the cell line model. In methacarn-fixed samples mRNA amount was also preserved, whereas in formalin and FineFIX-fixed samples it was notably lower when compared with the fresh frozen control. Alcohol-based fixatives are a good solution for long-term fixation of both cytologic and tissue samples by virtue of their time-independent effects on mRNA preservation. In fixed tissue samples, however, the potential effects of preanalytical tissue-related factors should be considered when performing mRNA quantitative analysis.

Lumboaortic and iliac lymphadenectomy: what is the role today?

PURPOSE: The aim of this study was to evaluate the roles of the lymphadenectomy in the surgical treatment of rectal cancer. METHODS: On the basis of our experience of 252 curative operations for rectal cancer, we analyze survival and recurrences in relation to the lymph node involvement and to the level of the lymph nodes where the metastases are located. All patients underwent a lymphadenectomy with high ligation of the inferior mesenteric artery and removal of the lumboaortic lymph nodes from the left renal vein to the aortic bifurcation. Pelvic lymphadenectomy was performed in 16 cases. RESULTS: Five-year survival was 70.6 percent in patients with no lymph node involvement, 68.2 percent in patients with pararectal lymph nodes N+, 25 percent in patients with involvement of intermediate lymph nodes, and 30 percent in patients with involvement of lumboaortic lymph nodes. In no case was there involvement of the hypogastric lymph nodes. On the basis of our experience and from results in the literature, we consider an upward extended lymphadenectomy with high ligation of the inferior mesenteric artery is warranted since it enables the tumor to be staged accurately and may lead to survival even in cases of advanced lymph node involvement.

Local excision for rectal cancer.

The aim of this retrospective study is to evaluate the results of local excision (LE) for rectal cancer for curative purposes.

Expression profiling of angiogenic genes for the characterisation of colorectal carcinoma.

The development of new blood and lymphatic vessels is a crucial event for cancer growth, metastatic spread and relapse after therapy. In this work, the expression levels of chemokines, angiogenic and angiostatic factors and their receptors were determined in paired mucosal and tumour samples of patients with colorectal carcinoma and correlated with clinical and histological parameters by advanced multivariate analyses. The most important predictors to discriminate between tumour and paired normal mucosa turned out to be the levels of expression of plexin-A1 and stromal cell-derived factor 1 (SDF-1), the former overexpressed and the latter downregulated in tumours. The levels of osteopontin and Tie-2 transcripts discriminated between the presence and absence of lymph node infiltration, the former overexpressed in the presence of infiltration whilst the latter providing a protective role. These results add support to the notion that the expression levels of selected genes involved in new blood and lymphatic vessel formation represent trustable biomarkers of tumour development and invasion and contribute to the identification of novel molecular classifiers for colorectal carcinoma.

Sentinel lymph node mapping in the management of colorectal cancer: preliminary report.

Aim and background The problem of understaging the lymph node status in colorectal cancer because of missed micrometastases led authors to investigate the role of sentinel node (SN) mapping also in colorectal malignancies. The aim of this study was to evaluate the feasibility of the technique and to correlate the results with some characteristics of the primary tumor. Methods Sentinel lymph node mapping was performed in 23 patients who underwent a standard lymphadenectomy for colorectal cancer. The vital dye Patent Blue had been injected into the peritumoral subserosa in vivo in 17 cases and ex vivo in seven, including one case where the in vivo method did not allow to identify the sentinel node. The nodes that took up the dye were removed and analyzed with standard hematoxylin-eosin staining in serial sections. Immunohistochemistry (AE1-AE3 cytokeratin markers) was performed in hematoxylin-eosin-negative nodes. SN status was related to the status of the other lymph nodes in the surgical specimen analyzed with the standard technique and to the following characteristics of the primary tumor: stage, grade and diameter. Results The in vivo technique allowed to identify the SN in 16/17 cases (94.1%), the ex vivo technique in 7/7. A total of 336 lymph nodes dissected from the surgical specimens was analyzed, with an average of 14.6 nodes per patient (range, 7-35). Of these nodes 58 were SNs, with an average of 2.5 nodes per patient (range, 1-8). In the 19 cases where the SN was tumor negative, the non-SNs were also negative (specificity: 100%), whereas in the four cases where the non-SNs were positive, in two cases the SN was positive and in two cases of pT3 rectal carcinoma the SN was negative (sensitivity: 50%). Immunohistochemistry did not modify the negative results of the standard hematoxylin-eosin evaluation. Conclusions The method used to identify the SN using vital dye proved to be easy to use both in vivo and ex vivo and allowed to identify the SN in all cases. The preliminary results indicate that there is a risk of false negative findings and therefore further studies are required to improve the sensitivity and the specificity of the technique and to evaluate the role of SN mapping in colorectal cancer management.

Migrating Acute Onset Abdominal Pain: Axial Torsion and Necrosis of a Giant Meckel’s Diverticulum Laparoscopically Resected

BackgroundTorsion is a rare complication of Meckel’s diverticulum. The case is reported of migrating, acute-onset abdominal pain due to axial torsion and necrosis of a Meckel’s diverticulum, diagnosed and treated laparoscopically.MethodsA 45-year-old woman presented with left upper quadrant abdominal tenderness and leukocytosis. On the following day, the tenderness had shifted to the right lower quadrant and was accompanied by rebound tenderness. Diagnostic laparoscopy revealed a twisted and necrotic giant Meckel’s diverticulum, which was completely laparoscopically managed.ResultsThe patient had an uneventful enhanced recovery. The laparoscopic approach avoided unnecessary scans and led to the easy diagnosis and surgical treatment without complications.ConclusionsMigrating, acute-onset non-specific abdominal pain may be due to a complication of a Meckel’s diverticulum, as in this case of torsion and necrosis.