H. J. Buhr

Quantitative real‐time RT‐PCR for detection of disseminated tumor cells in peripheral blood of patients with colorectal cancer using different mRNA markers

The detection of disseminated tumor cells in peripheral blood from colorectal cancer patients by RT‐PCR could be an attractive method for selecting patients for adjuvant therapy. We here report on real‐time RT‐PCR assays (LightCycler) to quantitate potential mRNA markers. We investigated specimens from colon carcinoma and normal colon mucosa tissues, cell lines, blood samples from 129 patients with colorectal cancer (all stages) and 58 reference blood samples (healthy donors, persons suffering from inflammatory bowel or infectious diseases). The expression profile in tissues showed high values for CEA and CK20, whereas in cell lines ProtM was predominant. All markers were detected in reference and patient blood samples (ProtM, 22, 17%; CEA, 84, 86%; CK20, 85, 88%). After quantitative analysis, the definition of cutoff values for each marker and the combination of markers, 13% of patients were judged to have elevated marker concentrations in their blood, from which only 6 had values significantly differing from cutoff value. There were no differences between stages of disease. In the case of 19 patients, investigated prior to and 1 week after surgery, 2 samples revealed a significant postoperative increase in CEA or CK20 mRNA concentration. In spite of high expression levels in tissues and cell lines, we were not able to differentiate satisfyingly mRNA markers originating from tumor cells and those from illegitimate transcription in hematopoetic cells in blood. We conclude that either copy numbers of analyzed markers in circulating tumor cells are not sufficient for detection or, more probably, peripheral blood is not a suitable compartment for detection of tumor cells in colorectal cancer.

Does glutamine reduce bacterial translocation? A study in two animal models with impaired gut barrier.

Abstract Failure of intestinal barrier function and subsequent translocation of bacteria from the gut are believed to play a decisive role in the development of systemic septic complications, for example, following major trauma or major abdominal surgery. This study evaluated: (a) the effect of glutamine on colonic microcirculation and electrophysiological parameters reflecting gut barrier function, (b) the translocation of live bacteria to extraintestinal organs, and (c) disease outcome in two animal models with impaired gut barrier function. Severe acute pancreatitis or colitis was induced in rats randomized for therapy with or without glutamine (0.5 g/kg daily). After 48 h one animal group was prepared for intravital microscopy of colonic capillary blood flow and electrophysiological measurement of gut permeability; another was killed after 96 h for histological and microbiological examination. In animals with pancreatitis, glutamine (Gln) supplementation significantly improved gut permeability, i.e., Gln increased colonic transmucosal resistance from 67±7 to 92±3 Ω/cm2 and decreased mannitol flux through the epithelium by 53%. Capillary blood flow in the colonic mucosa was improved by 25%. The prevalence of pancreatic infections was reduced from 86% in animals on standard parenteral nutrition to 33% in animals given the Gln-enriched diet (P<0.05); mortality decreased by 32%. In colitis, Gln had no significant effect on these parameters except for improving colonic capillary blood flow in colon segments not adjacent to the major injury site. Glutamine supplementation improves colonic capillary blood flow, stabilizes gut permeability, and reduces secondary pancreatic infections and mortality in severe rodent pancreatitis, but it is not helpful in colitis. This confirms previous reports that glutamine stabilizes gut barrier function only in certain diseases. Our experimental data strongly suggest that acute pancreatitis (rather than colitis) is one of the diseases with gut barrier dysfunction in which glutamine substitution may be helpful to reduce bacterial translocation and should therefore be tested in a controlled clinical trial.

Interstitial laser coagulation: Evaluation of the effect of normal liver blood perfusion and the application mode on lesion size

The effect of temporarily interrupted hepatic blood flow and multiple‐fiber application on necrosis volume in interstitial laser coagulation (ILC) was investigated.

Persistent Multiple Organ Microcirculatory Disorders in Severe Acute Pancreatitis: Experimental Findings and Clinical Implications

This study characterizes microcirculatory changes (capillary blood flow, capillary permeability, and leukocyte rolling) in the pancreas, colon, liver, and lungs at different stages of severe acute pancreatitis (AP) in a well-established rat model using intravital microscopy and computerized image analysis. The results demonstrate that microcirculatory disorders in severe AP are not confined to the pancreas but can also be found in the colon, liver, and lungs; that they extend beyond the early stage of AP and persist for 48 hr (and longer); and that they not only affect capillary blood flow but also involve prolonged changes of capillary permeability and leukocyte endothelial interaction. These findings may explain previous observations that therapeutic strategies aimed at enhancing microcirculation improve outcome in AP even if therapy is delayed and pancreatic necrosis can no longer be influenced. Since these systemic microcirculatory disturbances may contribute to AP-associated multiple organ dysfunction syndrome, further studies are warranted to evaluate whether improvement of microcirculation stabilizes organ function in AP and how long this may be effective after disease onset.

Does microcirculation play a role in the pathogenesis of inflammatory bowel diseases? Answers from intravital microscopic studies in animal models

Abstract The potential role of intestinal microcirculation for the development of inflammatory bowel diseases (IBD) has not been systematically investigated, mainly because of methodological problems. Using a well-established rodent model of IBD and intravital microscopy, the present study investigated whether (and when) gut microcirculation is disturbed in IBD, and whether microcirculatory disorders contribute to histological and functional alterations in the development of IBD. Colitis was induced by rectal injection of trinitrobenzene sulfonic acid. After 1, 3, and 15 days rats were laparotomized for intravital microscopic determination of mucosal colonic blood flow. In a second series it was examined whether enhancing colonic capillary blood flow by hemodilution therapy stabilizes colonic wall resistance and other electrophysiological parameters of gut permeability. Additional measurements involved hemodynamic monitoring and histological examinations. Colonic capillary blood flow was significantly decreased 3 days after colitis induction (1.8±0.05 vs. 2.6±0.04 nl/min in healthy control animals) when histology revealed signs of acute inflammation, and normal values after 15 days (2.4±0.06 nl/min) when chronic histological changes were evident. Hemodilution therapy enhanced colonic capillary blood flow in the initial stage (2.1±0.02 vs. 1.6±0.02 nl/min in saline-treated animals with trinitrobenzene sulfonic acid colitis) and improved gut resistance and electronic chlorid secretion (73±15 vs. 33±8 μA cm2). Histological alterations were not significantly attenuated. Impaired colonic capillary blood flow in the initial stage of experimental colitis and improved mucosal microcirculation with stabilized gut permeability suggests that the early microcirculatory disturbances precede chronic histological changes and influence functional alterations in the course of the disease. Research should be continued in this field because important mechanisms in the pathogenesis of IBD and potentially therapeutic (vasoactive) substances may otherwise be overlooked.

Expression of SIALYL-Lex antigen defined by MAb AM-3 is an independent prognostic marker in colorectal carcinoma patients

Expression of mucin‐bound sialyl‐Lex antigen during the progression of colorectal carcinoma and its potential prognostic value were analysed in sections of tumours from 182 patients with a documented follow‐up by immunohistochemistry using the monoclonal antibody (MAb) AM‐3. Two groups of colonic carcinomas with weak (n = 79) and strong (n = 103) sialyl‐Lex expression were discerned. The percentage of strongly expressing tumours increased with the progression of the disease (UICC stage I = 10%, stage II = 46%, stage III = 63%, stage IV = 68%, p < 0.0001). Seventy‐four percent of patients with carcinomas exhibiting a strong sialyl‐Lex expression but only 34% of patients with weak sialyl‐Lex expression died of the disease (p = 0.0026). In multivariate analysis, strong sialyl‐Lex expression increased the relative risk of cancer‐related death 3.8‐fold (95% CI = 1.8–7.9, p = 0.00034). The separate analyses of patients in UICC stage II (n = 56), III (n =5 9) and IV (n = 57) revealed that strong sialyl‐Lex expression was associated with a reduction of the 5‐year overall survival rate in UICC stage II (84% vs. 54%, p = 0.0013) and in stage III patients (86% vs. 35%, p = 0.0008) after curative resection but was not relevant in patients with distant metastases. In conclusion, the strong expression of sialyl‐Lex antigen defined by the MAb AM‐3 in colorectal carcinomas is an independent unfavourable prognostic factor after curative resection in stage II and III patients. The predictive power of the sialyl‐Lex expression may be helpful to define subgroups of patients at high risk for whom preventive adjuvant therapy can be selectively applied before the occurrence of detectable metastases. Int. J. Cancer 88:281–286, 2000.

Laser-induced thermotherapy for the treatment of liver metastasis: Correlation of gadolinium-DTPA-enhanced MRI with histomorphologic findings to determine criteria for follow-up monitoring

AbstractPurpose: To evaluate gadolinium (Gd)-diethylenetriamine-pentaacetic-acid (DTPA)-enhanced magnetic resonance imaging (MRI) for follow-up monitoring of laser-induced thermotherapy (LITT) and to determine a useful examination schedule. Methods: LITT of the liver was performed in 55 rabbits using a neodymium: yttrium-aluminum-garnet (Nd:YAG) laser (4-W power output, 840-s exposure time). Gd-DTPA MRI and histologic examinations were performed at different times (0–168 days). Results: Laser-induced lesions underwent regeneration and volume size reduction (69% after 168 days). The correlation coefficient (MR vs. macroscopic analysis) for the mean lesion diameter was r= 0.96. Histology of lesions comprised the four zones that correlated best with MRI findings. Coagulation necroses immediately after LITT was seen as an area of no enhancement on Gd-DTPA MRI. Circular enhancement was first seen 72–96 h after LITT, which was due to early mesenchymal proliferation. Conclusions: Gd-DTPA MRI is a good monitoring procedure for LITT. MRI should be performed 24 and 96 h after LITT.

Experimental models of acute pancreatitis: are they suitable for evaluating therapy?

Abstract Since randomized controlled studies of severe acute human pancreatitis can be performed only with restrictions, at least some aspects of innovative therapy concepts should first be clarified in animal experiments. In vitro trials are inadequate for this purpose since they cannot simulate the complex course of severe acute pancreatitis. Animal test results can be transferred to clinical practice if the results are based on trials with established models, standardized methods, and a study design imitating the clinical situation. This contribution discusses the demands on such an animal model of acute pancreatitis and a corresponding study protocol and presents models and protocols which meet these requirements. Concrete examples are presented to show that animal experiments are of great value under these conditions, especially in acute necrotizing pancreatitis. Further standardization of models, protocols, and monitoring should further improve future animal therapy studies at least to the extent that it is possible to select particularly promising substances, which should then be tested in randomized controlled trials.

Manometric analysis of anal sphincter damage after ileal pouch–anal anastomosis

Abstract A constant reduction in anal sphincter pressure follows an ileoanal pouch procedure for ulcerative colitis and familiar adenomatous polyposis. We analyzed whether this reduction is more likely due to neurogenic damage or to direct sphincter trauma. Three-dimensional vector volume manometry was performed in 75 patients prior to the ileoanal pouch procedure and 3 months thereafter. Resting pressure was significantly reduced from 83.5± 24.4 to 58.1±18.0 mmHg and squeezing pressure from 204.7±63.3 to 173.4±50.6 mmHg. Moreover, significant vector volume reductions were recorded postoperatively, and the asymmetry index increased significantly (resting: 11.5±4.1% to 18.4±7.4%; squeezing: 9.6±3.1 to 13.0 ± 6.7%). Functional anal sphincter length at the high-pressure zone remained unchanged. Thus, there was no local damage to proximal or distal anal sphincter segments, which suggests that the postoperative impairment of sphincter function is secondary to neurogenic rather than morphological damage.

SigmadivertikulitisOperationsindikation und -zeitpunkt

ZusammenfassungBei der chirurgischen Therapie der Kolondivertikelkrankheit sind OP-Indikation und -zeitpunkt vom Stadium der Erkrankung abhängig. Daher ist eine prätherapeutische, klinisch-pragmatische Stadieneinteilung die Voraussetzung für eine stadienadaptierte Therapie. Darüber hinaus sind Kenntnisse über den Spontanverlauf der Erkrankung, den Verlauf nach konservativer und operativer Therapie sowie über die individuellen Risikofaktoren für die Entwicklung eines komplizierten Verlaufs notwendig, um die richtige Indikation zur OP zu stellen. Bei der blanden Divertikulose und der unkomplizierten Divertikulitis ist keine Operation indiziert, die akute komplizierte Divertikulitis dagegen ist eine generelle OP-Indikation. Entscheidend für das Stellen der OP-Indikation ist daher eine möglichst exakte prätherapeutische Differenzierung zwischen komplizierter und unkomplizierter Divertikulitis. In Abhängigkeit von der Komplikationsart und dem klinischen Bild ist der OP-Zeitpunkt für die akute komplizierte Divertikulitis notfallmäßig oder frühelektiv nach initial konservativer und/oder interventioneller Therapie festzulegen. Bei der chronisch-rezidivierenden Divertikulitis ist ebenfalls eine OP indiziert. Als OP-Zeitpunkt ist die elektive Intervalloperation nach dem zweiten entzündlichen Schub anzustreben. Risikogruppen wie z. B. immunsupprimierte Patienten sollten bereits nach dem ersten entzündlichen Schub operiert werden.AbstractIndication and time for surgery of diverticular disease are determined by the stage of the disease. Clinically pragmatic pretreatment staging is thus a prerequisite for stage-adapted therapy. The correct indication for surgery is also based on knowledge of the spontaneous disease course, its course after conservative and operative therapy and the individual risk factors for complicated diverticular disease. Surgery is not indicated for bland diverticulosis or uncomplicated diverticulitis. It is generally indicated, however, for acute complicated diverticulitis. Decisive in establishing the indication for surgery is therefore the precise pretherapeutic differentiation of complicated and uncomplicated diverticulitis. Depending on the type of complication and the clinical appearance, the time for surgery of acute complicated diverticulitis is fixed on an emergency or early elective basis following initial conservative and/or interventional therapy. Chronically recurrent diverticulitis is likewise an indication for surgery. In terms of timing, an elective interval operation is best after the second inflammatory episode but should already be performed after the first one in risk groups, e.g. immunosuppressed patients.