Percutaneous fixation of acute scaphoid waist fractures: Long-term patient-reported functional outcomes and satisfaction at a mean of 11 years following surgery.

The aim of this study was to report the long-term functional outcomes and complication rates following early percutaneous fixation of acute fractures of the scaphoid. A trauma database was searched to identify all skeletally-mature patients with an undisplaced or minimally-displaced scaphoid waist fracture managed with early percutaneous retrograde screw fixation over a thirteen-year period from 1997-2010. Medical records were retrospectively reviewed, and complications documented. Long-term follow-up was by a questionnaire-based review. The Patient-Rated Wrist Evaluation (PRWE) was the primary outcome measure. Secondary outcomes included the Quick version of the Disability of the Arm, Shoulder and Hand score (QuickDASH), the EuroQol 5-dimensions score (EQ-5D-5L), and complications. During the study period 114 patients underwent this procedure. The mean age was 28 years (range 17-62) and 97 patients (85%) were male. The median time from injury to surgery was nine days (range 1-27). Twelve patients (11%) reported a complication, all of whom required repeat surgical intervention (six revision ORIF for non-union, five elective removal of hardware, one early revision fixation due to screw impingement). Long-term outcome data was available for 77 patients (68%) at mean follow-up of 11.4 years (range 6.4-19.8). The median PRWE was 0 (IQR 0-7.5), median QuickDASH 0 (IQR 0-4.5) and median EQ-5D-5L 1.0 (IQR 0.837-1.0). There were 97% (n = 74) patients satisfied with their outcome. Early percutaneous fixation of acute non-displaced or minimally displaced scaphoid fractures results in good long-term patient reported outcomes and health-related quality of life. Although comparable with previous studies, the overall surgical reintervention rate is notable and can result in inferior outcomes. LEVEL OF EVIDENCE: Therapeutic level III (Retrospective Cohort Study).

Does an evidence-based healthy relationships program for 9th graders show similar effects for 7th and 8th graders? Results from 57 schools randomized to intervention.

Integrating social and emotional learning (SEL) programming throughout curricula to support the development of healthy behaviors and prevent violence is critical for a comprehensive approach to school health. This study used a post-test comparison design to evaluate a healthy relationships program for eighth grade students that applies a SEL approach. The program was adapted from the Fourth R, an evidence-based program for ninth graders, but matches the curriculum and developmental context for eighth graders. Surveys were collected post-intervention from 1012 students within 57 schools randomized to intervention or control conditions. Multivariate multilevel analysis accounted for the nested nature of students within schools. There were significant group differences on three of four outcomes following intervention, including improved knowledge about violence, critical thinking around the impact of violence, and identification of more successful coping strategies. There was no group difference on general acceptance of violence. Overall, students learned relevant information and strategies and were able to apply that knowledge to demonstrate critical thinking, suggesting that adapting an evidence-based approach for use with younger students provided similar benefits. These findings build a case for 2 years of consecutive evidence-based healthy relationships programming in grades 8 and 9, consistent with best practice guidelines.

Clinicopathologic and prognostic significance of matrix metalloproteinases in rectal cancer.

BACKGROUND AND AIMS: The aim of this study was to determine the prognostic role of matrix metalloproteinases in rectal cancer. MATERIALS AND METHODS: Formalin-fixed and paraffin-embedded tissue sections of 94 rectal carcinomas were used for the immunohistochemical analysis of matrix metalloproteinases (MMP)-2, MMP-7, MT1-MMP, and tissue inhibitor of metalloproteinases (TIMP)-2. Inclusion criteria were sporadic rectal adenocarcinoma resected curatively (including total mesorectal excision), adjuvant radiochemotherapy in UICC stages II and III, and complete intra-institutional follow-up. Results of immunohistochemistry were correlated with clinical and histopathologic data from the prospective rectal cancer registry and prognosis. End points of the prognostic analysis were tumor progression caused by local and/or distant recurrence and 5-year survival (disease-free and overall). To assess prognostic significance, statistics included univariate and multivariate analysis (p<0.05 statistically significant). RESULTS: Of the 94 rectal carcinomas, 35% (33/94) showed an epithelial MMP-2 expression, 77% (72/94) were MMP-2 positive in the stroma. Fifty-four percent (51/94) were MMP-7 positive, and 47% (46/94) were positive for both MT1-MMP and TIMP-2. The stromal MMP-2 staining pattern was correlated with the depth of invasion (pT status, p=0.006) with MMP-7 (p=0.016) and TIMP-2 expression (p=0.036). Positive expression of MMP-2 in tumor epithelium was correlated with MMP-7 (p=0.027), MT1-MMP (p=0.036), and TIMP-2 expression (p<0.0001). A positive staining pattern of MMP-7 was significantly correlated with depth of invasion and TIMP-2 (p<0.01). The positive staining pattern of MT1-MMP was correlated with epithelial MMP-2 (p=0.036), MMP-7 (p=0.004), and TIMP-2 expression (p=0.002). TIMP-2 immunoreactivity correlated with depth of invasion (p=0.013), epithelial MMP-2 (p<0.001), stromal MMP-2 (p=0.036), MMP-7 (p<0.001), and MT1-MMP (p=0.002). Neither pattern correlated with age, gender, tumor stage (UICC), grading, preoperative serum carcinoembryonic antigen (CEA) level, or nodal status (p>0.05). Within a mean follow-up of 46 months, tumor progression, caused by either local recurrence or distant metastasis, occurred in 14 patients (15.4%). There was no significant association between the MMP expression and the incidence of local and/or distant recurrence. In terms of survival, preoperative CEA level (disease-free 5-year survival 46% with increased CEA vs 70% with normal CEA, p=0.01; overall 5-year survival 43 vs 74%, p<0.01) and UICC stage were the only factors to be significantly related to 5-year survival by univariate analysis, whereas the metalloproteinases failed to show a significant association. In multivariate analysis, CEA and UICC stage were not identified as independent factors predictive of survival. CONCLUSION: MMP-2, MMP-7, MT1-MMP, and TIMP-2 do not appear to be significant predictors of prognosis in a homogenous collective of curatively resected rectal adenocarcinomas.

Metronomic trofosfamide inhibits progression of human lung cancer xenografts by exerting anti-angiogenic effects.

BACKGROUND: Recent studies of conventional chemotherapeutic drugs administered in metronomic therapy schedules showed remarkable inhibitory effects on tumor angiogenesis. Subsequent and prolonged tumor regression was achieved moreover by circumventing acquired drug resistance. In this study, metronomic and conventional trofosfamide were compared on human NSCLC xenograft "LX-1." MATERIALS AND METHODS: In vitro cytotoxicity of trofosfamide on tumor and human umbilical cord endothelial cells was determined under normoxic and hypoxic conditions. Additionally fractions and duration of cell cycles were analyzed by flow cytometry. In vivo LX-1 xenotransplanted nude mice were treated with trofosfamide in conventional and metronomic schedules (i.p./p.o.). Tumor sections were evaluated for microvessel density (MVD), relative growth fraction and apoptosis. RESULTS: In contrast to the rapid growth of conventionally treated lung cancer, long lasting tumor growth retardation over the total treatment period was achieved with metronomic treatment. While growth fraction and apoptotic rate of LX-1 cells remained unchanged, the MVD was significantly reduced (50%). CONCLUSION: Our results show advantages of a metronomic trofosfamide schedule compared to a conventional bolus therapy mainly due to inhibition of angiogenesis. In vitro data show that this mechanism works under normoxic and hypoxic conditions and suggest that this is in part a direct cytotoxic effect on endothelial cells.

Influence of thymidylate synthase and p53 protein expression on clinical outcome in patients with colorectal cancer.

AIMS: Thymidylate synthase (TS) and tumor suppressor p53 are two proteins with an influence on tumor resistance to radio-chemotherapy that is well known. For this reason we tested the effect of TS and p53 expression on clinical outcome (tumor recurrence and survival) in patients after curative tumor resection, especially in patients who received adjuvant radio-chemotherapy. PATIENTS AND METHODS: A total of 120 patients with colorectal cancer were included in the study. A curative resection was possible in 83 patients, and 30 of this group received adjuvant therapy. For the immunohistochemical staining of tumor specimens, monoclonal antibody (mAb) TS 106 against TS and mAb DO-1 against p53 protein were used. TS positivity was defined as a moderate to high staining intensity in the cytoplasma of cells and p53 positivity as nuclear staining of tumor cells in >10% of these cells. RESULTS: Thymidylate synthase immunoreactivity was found in 59% of all cases and p53 staining in 51%. No relation between clinicopathological features and p53 expression was found in contrast to TS expression, where a highly significant association of TS-positive cases with tumor invasion (pT) was observed. Curatively resected patients with a TS-positive tumor developed tumor recurrence/distant metastases significantly more often than TS negative tumors. The same result was found when comparing p53-positive with p53-negative tumors and TS+/p53+ with TS-/p53- tumors. TS expression was highly significantly associated with poor survival and was the strongest independent prognostic factor in multivariate analysis, followed by lymph node status. CONCLUSION: Thymidylate synthase expression seems to be an independent prognostic factor and a possible predictor of tumor recurrence in patients with colorectal cancer.

Colorectal carcinomas with high MIB-1 labelling indices but low pKi67 mRNA levels correlate with better prognostic outcome.

AIMS: Antibodies specific to the proliferation-associated protein pKi67 (e.g. Ki67, MIB-1) are routinely used in oncology to assess the proliferation index of tumour cells. In untransformed cells the amount of pKi67 present at any time of the cell cycle is strictly regulated. To achieve a better understanding of expression and regulation of this protein in tumour cells, we investigated both pKi67 mRNA and protein expression in routinely fixed and embedded tissue of colorectal carcinoma. METHODS AND RESULTS: We determined a median pKi67 specific in-situ hybridization labelling index of 42% (9-79%) and a median Ki67 index (MIB-1 labelling index) of 59% (26-94%) in 47 resected colorectal adenocarcinomas of different stages and grades. In 32 cases expression of pKi67 mRNA and protein correlated well but we observed a significant difference between both values in 15 tumours. In these cases more than 30% of the cells expressed the protein but not the mRNA of pKi67, possibly due to cell cycle arrest. Patients belonging to this group had a significantly (P < 0.012) better prognosis. CONCLUSIONS: Tumours with a high pKi67 protein level but low mRNA expression are likely to proliferate more slowly than calculated on the basis of their Ki67 staining index, which possibly explains the patients' improved outcome.

Prognostic significance of p21 and p27 protein, apoptosis, clinical and histologic factors in rectal cancer without lymph node metastases.

The aim of this study was to evaluate the prognostic role of clinical and histopathologic factors, cell-cycle regulator proteins (p21(Waf1/Cip1), p27(Kip1)), and apoptotic index in lymph node-negative rectal cancer. Formalin-fixed, paraffin-embedded tissue samples of 97 rectal carcinomas (UICC stages I and II) resected curatively within five years were used. Immunohistochemical analysis of protein expression was performed by monoclonal antibodies: p21 (clone SX118), p27 (clone SX53G8). Apoptosis was assessed by the TUNEL method. Clinical, surgical, histopathologic, and follow-up data were prospectively recorded in a computerized registry. To assess prognostic significance (end points: metachronous distant metastases, 5-year disease-free and overall survival), statistics included univariate and multivariate analysis (p < 0.05 statistically significant). Of the 97 rectal carcinomas without lymph node metastases, 46.4% (45/97) were p21-positive, 49.5% were p27-positive (48/97), whereas 27.8% (27/97) showed a high apoptotic index. Within a median follow-up of 54 months, 4 patients developed local recurrence (4.1%). Distant metastases occurred in 12 patients (12.4%). Univariate analysis showed that gender, UICC stage, p21 and p27 were significantly associated with the incidence of distant metastases (p < 0.05). UICC stage and p21 were the only factors to be significantly related to 5-year disease-free survival by univariate analysis (p < 0.05). Only UICC stage was significantly related to 5-year overall survival (p < 0.05). The apoptotic index was correlated neither to recurrence nor to survival (p > 0.05). Multivariate analysis demonstrated that gender, UICC stage and p21 were independently related to the incidence of distant metastases; however, UICC stage was the only independent factor predictive of 5-year disease-free survival and overall survival (p < 0.05).

[Influence of resterilized polypropylen meshes on growth of human fibroblasts--an experimental in vitro study]. / Einfluss resterilisierter Kunststoffnetze aus Polypropylen auf das Wachstum humaner Fibroblasten--Eine experimentelle in vitro Studie.

INTRODUCTION: We investigated the influence of resterilized polypropylen meshes (Prolene(R)) on proliferation and apoptosis of human fibroblasts in an experimental in vitro study. METHOD: Human fibroblasts were seeded into six-well culture dishes in a density of 3 x 10 (4) cells/well. After resterilization of meshes (steam autoclave, 121 degrees C, 20 min.) according to the manufacturer's recommendations (Ethicon, Norderstedt) square sheets of 2 x 2 cm were incubated with fibroblasts over a period of 6, 12, 18, 24, 30, 36, 42 and 48 h. Preparations of fibroblasts with non-resterilized meshes and without meshes served as controls. Proliferation index and apoptotic index were estimated by flow cytometry after cell staining with an FITC-conjugated antibody against the Ki-67 antigen or with FITC-conjugated Annexin-V and propidium jodide, respectively. RESULTS: A significant reduction of the proliferation index from 86 % to 42 % was found after 48 h incubation of cells with resterilized meshes, whereas only a slight decrease was found in the group with non-resterilized meshes (75 %) and in controls without meshes (80 %). Apoptotic index increased significantly from 2 % to 48 % after 48 h incubation with resterilized meshes in comparison to both control groups, where only a slight increase could be observed: non-resterilized meshes to 19 % and without meshes to 10 %. CONCLUSION: Resterilized meshes inhibit growth of human fibroblats in vitro significantly, demonstrated by a reduced proliferative activity and an increased apoptotic index. This could be caused by a release of toxic substances from the meshes, which have a negative influence on cell growth. Therefore, resterilization cannot be recommended.

p21, p27, cyclin D1, and p53 in rectal cancer: immunohistology with prognostic significance?

BACKGROUND AND AIMS: This study examined the prognostic value of the cyclin-dependent kinase inhibitors p21(Waf1/Cip1) and p27Kip1, and the cell cycle regulating proteins cyclin D1 and p53 after curative surgery for rectal cancer. PATIENTS AND METHODS: Formalin-fixed, paraffin-embedded tissue samples of 160 rectal carcinomas resected curatively within a 5-year period were used. Immunohistochemical analysis used monoclonal antibodies p21(Waf1/Cip1) (clone SX118), p27Kip1 (clone SX53G8), cyclin D1 (clone DCS-6), and p53 (DO-1). Positive nuclear protein expression was assessed at the 10% level. Results of immunohistochemistry were studied for correlation with clinical and histopathological data of the prospective tumor registry including recurrence and patient survival. RESULTS: Of the 160 rectal carcinomas 36% were p21(Waf1/Cip1) positive, 44% p27Kip1 positive, 48% cyclin D1 positive, and 39% p53 positive. The p21(Waf1/Cip1) staining pattern was correlated with p27Kip1 and p53 expression and with UICC stage and lymph node status. p53 status was not correlated to any clinical or histopathological variable. p27Kip1 expression was associated with tumor size and cyclin D1 expression. Tumor progression caused by local and distant recurrence occurred in 20%. p21(Waf1/Cip1), p27Kip1, and p53 were strong predictors of recurrence. p21(Waf1/Cip1) and p53 but not p27Kip1 were independently correlated with disease-free survival. UICC stage was independently related to both recurrence and survival. The best prognosis was in p21(Waf1/Cip1) positive and p53 negative rectal carcinomas. CONCLUSIONS: Reflecting tumor biology by immunohistochemical assessment of cell cycle regulators, p21(Waf1/Cip1) and p53 were independently predictive of prognosis in rectal cancer, and p27Kip1 was independently related to recurrence. However, cyclin D1 had no independent relationship to prognosis. Clinically, UICC stage was a strong predictor of prognosis after curative surgery for rectal cancer.

[Polypropylene synthetic mesh modifies growth of human cells in vitro. An experimental study]. / Kunststoffnetze aus Polypropylen beeinflussen das Wachstum humaner Zellen in vitro. Eine experimentelle Studie.

INTRODUCTION: The aim of our study was to investigate the influence of polypropylene meshes on the proliferation and apoptosis of human cell cultures in vitro. METHODS: Human fibroblasts and HeLa cells were incubated in different densities (10(4), 3.10(4), 10(5) cells/well) together with polypropylene meshes (Prolene, 2 x 2 cm) in six-well culture dishes over a 48-h period. Cells without meshes served as controls. Cells were spun on slides and stained with the monoclonal antibody MIB-1. To calculate the proliferation indices, stained nuclei were counted. Apoptotic indices were determined by flow cytometric analysis, using FITC-conjugated Annexin-V and propidiumjodide for staining. RESULTS: Fibroblasts showed only a slight reduction of the proliferation index (PI) from 64% (controls) to 60% (meshes). Increasing cell density leads to a decrease in the PI of both groups. The PI of HeLa cells was similar in mesh groups and controls and independent of cell density. The apoptotic index (AI) of fibroblasts was significantly higher in the mesh group (3.7%) in comparison with the controls (1.9%). The same was observed for HeLa cells (AI mesh group: 4.5%, AI controls: 1.2%). Furthermore, an increase of AI was found with increasing cell density in both cell lines. CONCLUSION: Whereas meshes did not influence the proliferation of the cell lines examined, they seem to have a marked influence on apoptosis, as a significant increase of AI was observed in the mesh group in contrast to controls.

Assessment of proliferative activity in colorectal carcinomas by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR).

The monoclonal antibody Ki-67 and the isospecific monoclonal antibody MIB-1 are routinely used in oncology to assess the proliferation index of tumor cells. A more objective and sensitive method is the determination of the of Ki-67 protein-specific mRNA by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). In 25 resected colorectal adenocarcinomas of different stages and grades we determined between 0.2 and 4.4 amol (10(-18) mol) Ki-67 protein-specific mRNA per microgram total RNA (median = 0.88 amol). The corresponding Ki-67 indices (expressing the percentage of Ki-67/MIB-I positive tumor cells) ranged from 41 to 81% (median = 61%). We found a good correlation between Ki-67 index and mRNA expression (r = 0.75), a significant correlation between both data and tumor stage (primary tumor, regional nodes, metastasis [pTNM] staging classification) (p < 0.001), but not between both data and tumor grade. Both Ki-67 indices (p = 0.05) and mRNA levels (p = 0.014) correlated significantly to the patients' survival. These results demonstrate that the Ki-67 protein-specific quantitative RT-PCR is a useful method for the characterization of tumor cell proliferation.

Prognostic significance of free gastrointestinal tumor cells in peritoneal lavage detected by immunocytochemistry and polymerase chain reaction.

AIMS: The aim of our study was to identify tumor cells in peritoneal lavage comparatively with immunocytochemistry (ICC) and half-nested reverse transcriptase-polymerase chain reaction (RT-PCR) using carcinoembryonic antigen (CEA) as marker and to evaluate their prognostic significance. PATIENTS AND METHODS: In 75 patients who underwent surgery for a carcinoma of the colorectum (n=49), stomach (n=17) or pancreas (n=9) and 13 patients with an abdominal aortic aneurysm (control group) the abdomen was irrigated with saline solution immediately after laparotomy. Cells were separated by Ficoll-density centrifugation and divided into 2 equal volumes for ICC and RT-PCR. For ICC cells were spun onto slides by cytospin centrifugation and stained with a monoclonal antibody (mab) against CEA using the APAAP method. For RT-PCR total RNA was extracted from the cells, transcribed into cDNA and amplified with CEA-specific primers. Lavages of 13 patients with an abdominal aortic aneurysm and blood samples of 6 healthy donors served as controls. RESULTS: Immunostained tumor cells were found in peritoneal lavage in 23% (17/75) of all patients, whereas 63% (47/75) of patients gave a positive result by RT-PCR analysis. In the control group (n=13) no patient presented with tumor cells in ICC, however 5 of 13 (38%) showed amplified CEA-mRNA by RT-PCR, and so did one of six blood samples. Using ICC technique, we found significant correlations between detection rates and pT-, pN-, pM-categories as well as tumor stage. On the contrary, by RT-PCR significant correlations were observed only between pT- and pM-categories and detection rates. Detection of tumor cells in peritoneal lavage with both techniques was associated with poor prognosis. Moreover, these tumor cells are an independent prognostic factor and may have an influence on the development of peritoneal carcinomatosis. CONCLUSION: ICC is a useful method for detection of tumor cells in peritoneal lavage. In contrast, half-nested RT-PCR cannot be recommended, as the detection rates are unproportionally high, obviously as a result of CEA-mRNA expression in nontumor cells.

p53 autoantibodies in sera of patients with a colorectal cancer and their association to p53 protein concentration and p53 immunohistochemistry in tumor tissue.

We evaluated p53 autoantibodies (p53-Ab) as a preoperative tumor marker and as a prognosis marker. We also investigated whether p53-Ab production is dependent on p53 protein overexpression in tumor tissue or on tumor volume. Serum samples of patients with a colorectal cancer (n = 130) and of healthy controls (n = 44) were examined for p53-Ab using an ELISA kit. P53 protein expression in tumor tissue was demonstrated immunohistochemically and quantified by ELISA. Tumor volume was calculated and patients' survival computed using the Kaplan-Meier method. p53-Ab were detected in the serum from 15% of patients; all controls were negative. There was a significant correlation between p53-Ab production and positive immunostaining or p53 protein concentration in tumor tissue. p53-Ab were detected at a higher percentage of patients with a tumor volume of 10 cm3 or greater than in those with a smaller tumor. No difference in patients' prognosis was found between the p53-Ab positive and negative groups. Because of their low sensitivity (15%) p53-Ab are not suitable as a preoperative tumor marker. However, their high specificity (100%) and their potential for early diagnosis of a tumor relapse makes them valuable for postoperative monitoring during follow-up in p53-Ab positive patients. Furthermore, their detection can be used as a simple serological test for early detection of p53 alterations.

Vascular endothelial growth factor (VEGF)--a valuable serum tumour marker in patients with colorectal cancer?

INTRODUCTION: Neo-angiogenesis, of great importance for tumour growth and nutrition, is preferentially mediated by the cytokine vascular endothelial growth factor (VEGF), which has a direct effect on vascular endothelial cell proliferation and migration. This study was designed to clarify whether VEGF is a suitable tumour marker in sera of patients with a colorectal cancer, and whether VEGF concentrations in sera and tumour tissues are correlated with tumour extension (pTNM) and especially with tumour volume or size. Furthermore, the influence of VEGF levels on patients >> prognosis was examined. METHODS: VEGF serum concentrations of 122 patients with colorectal cancer and 65 controls were determined with an ELISA kit. Additionally, VEGF concentrations of tumour and normal tissue were measured in 38 patients using the same ELISA. RESULTS: Our results demonstrate that VEGF is not a suitable diagnostic tumour marker in patients with colorectal cancer due to its low sensitivity (36%). However, a combination of the serum tumour markers CEA and VEGF can significantly increase the pre-operative diagnostic sensitivity to 62%. VEGF serum levels differed significantly between patients (mean 438 pg/ml) and controls (mean 203 pg/ml), and also between tumour and normal tissue (984 vs 89 pg/mg protein). Serum concentration showed a significant correlation to tumour volume and size. Patients with VEGF serum levels greater than cut-off had a poorer prognosis than those less than or equal to cut-off. For this reason VEGF could be used as a predictor of patients >> outcome.

Suppression of cell division by pKi-67 antisense-RNA and recombinant protein.

The human antigen defined by the monoclonal antibody Ki-67 (pKi-67) is a human nuclear protein strongly associated with cell proliferation and found in all tissues studied. It is widely used as a marker of proliferating cells, yet its function is unknown. To investigate its function we suppressed pKi-67 expression by antisense RNA and overexpressed a partial structure of pKi-67 in HeLa cells. A BrdU-incorporation assay showed a significant decrease in DNA synthesis after antisense inhibition. Cell cycle analysis indicated a higher proportion of cells in G1 phase and a lower proportion of cells in S phase while the number of G(2)/M phase cells remained constant. Overexpression of a recombinant protein encoding three of the repetitive elements from exon 13 of pKi-67 had a similar effect to that obtained by antisense inhibition. The similarity of the effect of expressing 'Ki-67 repeats' and pKi-67 antisense RNA could be explained by a negative effect on the folding of the endogenous protein in the endoplasmatic reticulum. Furthermore excessive self-association of pKi-67 via the repeat structure could inhibit its nuclear transport, preventing it from getting to its presumptive site of action. We conclude that the Ki-67 protein has an important role in the regulation of the cell cycle, which is mediated in part by its repetitive elements.

Apoptosis in rectal cancer: prognostic significance in comparison with clinical histopathologic, and immunohistochemical variables.

PURPOSE: The aim of this study was to evaluate the prognostic value of the apoptotic index for recurrence and disease-free survival after curative surgery for rectal cancer, particularly in relation to clinicopathologic variables, p53- and bcl-2 expression. METHODS: Formalin-fixed, paraffin-embedded tissue samples of rectal carcinomas resected curatively within a five-year period were used (N = 160). Apoptotic cells with fragmented DNA were detected by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphatase-biotin nick-end-labeling method. The ratio of apoptotic tumor cells (in percent) was classified into low apoptotic index (less than 10 percent) and high apoptotic index (10 percent or more). Immunohistochemical analysis was performed using monoclonal antibodies (DO-1 for p53 and clone 124 for bcl-2). Statistics included univariate and multivariate analysis, and survival was calculated using the Kaplan-Meier method. RESULTS: Seventy-five percent of tumors showed a low apoptotic index, and 25 percent had a high apoptotic index. No correlation was found between apoptotic index and International Union Against Cancer stage (P > 0.05). However, significant correlations were documented with histologic differentiation (mean apoptotic index, 5.74 percent in moderately vs. 3.98 percent in poorly differentiated carcinomas; P = 0.0173), lymph node involvement (mean apoptotic index, 6.11 percent in pN1 vs. 3.72 percent in pN2; P = 0.0074), p53 status (mean apoptotic index, 6.26 percent in p53- vs. 4.42 percent in p53+; P = 0.0085), and bcl-2 expression (mean apoptotic index, 5.13 percent in bcl-2- vs. 6.51 percent in bcl-2+; P = 0.0418). Tumors of the lower rectum had a lower apoptotic index than those of the upper rectum (P = 0.0277). Neither univariate nor multivariate analysis assessed apoptotic index as predictor of prognosis: Recurrence rates did not differ between tumors related to apoptotic index (22 percent with low apoptotic index vs. 15 percent with high apoptotic index; P > 0.05), and no significant differences were found regarding survival (P > 0.05). On multivariate analysis, International Union Against Cancer stage (P = 0.0002), p53 (P = 0.0002), gender (P = 0.0136), and bcl-2 (P = 0.0243) were independent predictors of recurrence. These variables, except for bcl-2, were also independently related to disease-free survival. CONCLUSIONS: Reflecting tumor biology, apoptotic index as single variable showed no prognostic significance, whereas p53 was an independent predictor for both recurrence and survival, and bcl-2 was independently related to recurrence, but not to survival. Clinically, International Union Against Cancer stage and gender were independent prognostic factors after curative surgery for rectal cancer.

p53 and Bcl-2 as significant predictors of recurrence and survival in rectal cancer.

The aim of this study was to evaluate the prognostic value of p53 nuclear accumulation and Bcl-2 expression after curative surgery for rectal cancer. Immunohistochemistry was performed using monoclonal antibodies (MAb) (DO-1 for p53; anti-human Bcl-2 MAb, clone 124, for Bcl-2) on formalin-fixed, paraffin-embedded tissues of 160 rectal carcinomas (UICC stages I-III), and results were compared with data from the prospective registry of rectal cancer by univariate and multivariate logistic regression model focusing specifically on recurrence. Survival was calculated by the Kaplan-Meier method and proportional hazards model. p53 nuclear accumulation was documented in 39% (n=63) of tumours and was associated with a higher incidence of tumour progression (local or distant recurrence) and poorer disease-free survival (P<0.0001). Bcl-2 expression was detected in 29% (n=47), and was associated with longer disease-free survival and lower incidence of recurrence (P<0.0086). Multivariate logistic regression analysis demonstrated that gender (P=0.0136), UICC stage (P=0.0002), p53 expression (P=0.0002) and Bcl-2 expression (P=0. 0243) were independent factors predictive of recurrence. The proportional hazards model identified p53 (P=0.0009), UICC stage (P=0.0480), gender (P=0.0049), but not Bcl-2 (P=0.1503), as independently related to disease-free survival. Looking at the p53/Bcl-2 subgroups, the poorest prognosis was observed in the p53+/Bcl-2- subgroup, whereas patients whose tumours were p53-/Bcl-2+ had the best prognosis (P<0.0001). Immunohistochemical assessment of both p53 and Bcl-2 status may be valuable in predicting recurrence and survival after curative surgery for rectal cancer. Therefore, they play a role as prognostic factors in rectal cancer. p53 is a stronger predictor of prognosis than Bcl-2.

[Ulcerative colitis-associated colorectal carcinoma. DNA ploidy as indicator of impending malignant transformation?]. / Das Colitis ulcerosa-assoziierte kolorektale Karzinom. DNS-Ploidie als Indikator einer drohenden malignen Transformation?

The onset of a malignant transformation in long-standing ulcerative colitis is difficult to predict. The value of the clinical and histomorphological parameters in current use is limited. It was thus aim of the present study to investigate the value of DNA-ploidy for the early detection of a malignant transformation in long-standing ulcerative colitis. This retrospective study comprised 20 patients with long-standing ulcerative colitis. The average observation time was 7.3 years (range: four to twelve years). All patients took part in a surveillance program and had between four and seven colonoscopies within a minimum period of time of five years. At these instances mucosal biopsies were taken in a standardized manner at eight different locations throughout the colon. These paraffin-embedded specimens (n = 542) were analyzed histomorphologically and DNA-cytometrically. During the observation time five patients developed an ulcerative colitis-associated colorectal carcinoma (UCA). In these patients epithelial dysplasias were not more common than in the remaining 15 cases. The vast majority of the specimen of the patients with UCA showed distinct DNA-cytometrical alterations, i.e. they were aneuploid. Such aneuploid mucosal cell populations were distributed over the whole colon, irrespectively of the later site of the carcinoma. These aneuploid lesions were found in one case eleven years, in an average seven years prior to the final diagnosis of a UCA. In contrast, the colon epithelium of the patients without UCA showed only proliferative-diploid DNA-distribution patterns during the observation time. In summary, affected patients had multiple highly aneuploid lesions of the colon mucosa at an average of seven years prior to the final diagnosis of UCA. These lesions came from macroscopically chronic inflamed tissue, and where histomorphologically without signs of dysplastic transformation. DNA-cytometrical investigations could thus be of additional predictive value for the individual risk assessment as regards an impending malignant transformation.

Expression of p53 and mdm2 mRNA and protein in colorectal carcinomas.

The aim of our study was to investigate the expression of p53 and mdm2 mRNA and protein in colorectal adenocarcinoma. For the detection of mRNA, 60 fresh frozen human tumour samples and 12 samples of corresponding normal tissue were examined. After total RNA extraction, reverse transcription (RT) was performed followed by cDNA amplification with specific primers using RT-polymerase chain reaction (PCR). Immunohistochemical detection of protein was examined in 81 formalin-fixed and paraffin-embedded human tumour specimens as well as 15 samples of adjacent normal colorectal tissue. p53 mRNA was detected in 80% (48/60) of the tumours and in 67% (8/12) of normal tissue samples; 87% (52/60) of tumours had mdm2 mRNA in contrast to only 17% (2/12) of normal tissue specimens. Nuclear p53 protein expression was observed in 52% (42/81) of the tumour specimens and in none of the 15 normal specimens, whereas mdm2 protein was found in the nucleus (31%, 25/81) and also in the cytoplasm (86%, 70/81) of tumour samples. In normal tissue, mdm2 protein expression was only observed in the cytoplasm (13%, 2/15) and not in the nucleus. There was a significant correlation between coexpression of p53 and mdm2 protein and the occurrence of lymph node metastases (P = 0.03) as well as between p53 protein expression and the occurrence of distant metastases (P = 0.007). Additionally, significant associations were found between p53 mRNA and p53 protein, p53 mRNA and mdm2 mRNA or protein, and also between mdm2 mRNA and mdm2 protein expression, supporting the existence of a regulatory mechanism involving p53 and mdm2.