Background Precise preoperative localization is essential for focussed parathyroidectomy. The imaging standard consists of cervical ultrasonography (cUS) and 99m Tc-MIBI-SPECT (MIBI-SPECT). 11 C-methionine positron emission tomography/computed tomography (Met-PET/CT) is a promising method for localizing parathyroid adenomas. The objective of our study was to elucidate whether additional Met-PET/CT increases the rate of focussed parathyroidectomy. Methods Fourteen patients with primary hyperparathyroidism (HPT) and three patients with tertiary HPT underwent cUS and MIBI-SPECT. Met-PET/CT was carried out in patients with negative MIBI results. Subsequent surgical strategy was adapted according to imaging results. Results cUS localized a single parathyroid adenoma in 10/17 patients (59 %), while MIBI-SPECT/CT identified 11/17 single adenomas (65 %). In the remaining six patients, Met-PET/CT identified five single adenomas. This step-up approach correctly identified single adenomas in 16/17 patients (94 %). Conclusion Met-PET/CT raises the rate of correctly localized single parathyroid adenomas in patients with negative cUS and MIBI-SPECT/CT and increases the number of focussed surgical approaches.

Serum calcium has been suggested to be a surrogate marker for intraoperative monitoring during a parathyroidectomy (PTX) for primary hyperparathyroidism (pHPT). The objective of the present prospective study was to investigate whether serum calcium can be used as an alternative for parathyroid hormone intraoperative monitoring.

BACKGROUND:In the case of hepatocellular carcinoma (HCC), underlying liver pathology may not only determine the feasibility of surgery but may also affect the postsurgical outcome. We report our experience after curative liver resection for HCC in patients with normal liver, liver fibrosis, and liver cirrhosis.METHODS:A total of 72 patients after liver resection with curative intention were analyzed. Histopathologic findings of tumor-unaffected liver tissue were used for retrospective classification: group A (normal liver); group B (liver fibrosis); group C (liver cirrhosis). The groups were compared for differences in short-term surgical results, total survival, and recurrence-free survival.RESULTS:The rate of major complications was 34.7% and did not significantly differ among groups. The overall perioperative mortality rate was 9.7%, with one patient dying in group A and three patients dying in each of the other two groups. Including perioperative mortality, the median overall survival for the whole group was 37.3 months (95% confidence interval 29.3-45.2 months). The respective 1-, 2-, and 5-year survival rates for group A (n = 21) were 86%, 71%, and 50% and for group C (n = 24) 62%, 50%, and 17%. The overall survival of group B (n = 27) was intermediate (log-rank, P = 0.032). The respective recurrence-free survival rates were 76%, 42%, and 20% for group A and 39%, 13%, and 4% for group C, with group B being intermediate (log-rank, P = 0.016).CONCLUSIONS:Our data demonstrate that liver resection in the presence of compensated liver cirrhosis is feasible but associated with a significantly impaired prognosis for overall and recurrence-free survival. The management of cirrhotic patients with compensated liver function and HCC therefore also requires the opportunity for transplantation.

Background: Metastasis formation is the leading cause of death among colon cancer patients. We established a new in-situ model of in vivo microscopy of the lung to analyse initiating events of metastatic tumor cell adhesion within this typical metastatic target of colon cancer.Methods: Anaesthetized CD rats were mechanically ventilated and 106 human HT-29LMM and T84 colon cancer cells were injected intracardially as single cell suspensions. Quantitative in vivo microscopy of the lung was performed in 10 minute intervals for a total of 40 minutes beginning with the time of injection.Results: After vehicle treatment of HT-29LMM controls 15.2 +/- 5.3; 14.2 +/- 7.5; 11.4 +/- 5.5; and 15.4 +/- 6.5 cells/ 20 microscopic fields were found adherent within the pulmonary microvasculature in each 10 minute interval. Similar numbers were found after injection of the lung metastasis derived T84 cell line and after treatment of HT-29LMM with unspecific mouse control-IgG. Subsequently, HT-29LMM cells were treated with function blocking antibodies against beta 1-, beta 4-, and alpha v-integrins wich also did not impair tumor cell adhesion in the lung. In contrast, after hydrolization of sialylated glycoproteins on the cells' surface by neuraminidase, we observed impairment of tumor cell adhesion by more than 50% (p < 0.05). The same degree of impairment was achieved by inhibition of P- and L-selectins via animal treatment with fucoidan (p < 0.05) and also by inhibition of the Thomson-Friedenreich (TF)-antigen (p < 0.05).Conclusions: These results demonstrate that the initial colon cancer cell adhesion in the capillaries of the lung is predominantly mediated by tumor cell - endothelial cell interactions, possibly supported by platelets. In contrast to reports of earlier studies that metastatic tumor cell adhesion occurs through integrin mediated binding of extracellular matrix proteins in liver, in the lung, the continuously lined endothelium appears to be specifically targeted by circulating tumor cells.

Recent studies have demonstrated that the chemokine receptor CXCR4 plays a crucial role in organ-specific metastasis formation. Although a variety of studies showed the expression of chemokine receptors, in particular, CXCR4, by gastrointestinal tumors, the precise mechanisms of chemokine receptor-mediated homing of cancer cells to specific sites of metastasis remained elusive. Here, we used liver metastatic human HEP-G2 hepatoma and HT-29LMM colon cancer cells expressing functional CXCR4 to dissect the metastatic cascade by intravital fluorescence microscopy. Immunohistochemistry revealed that the CXCR4 ligand CXCL12 is expressed by endothelial cells and likely Kupffer cells lining the liver sinusoids. Tumor cell adhesion and extravasation in vivo was quantitatively analyzed using intravital fluorescence microscopy. Treatment of cells with an anti-CXCR4 antibody did not affect cell adhesion but significantly impaired tumor cell extravasation (HEP-G2; isotype control: 22.3% +/- 4.3% vs anti-CXCR4: 6.0% +/- 5.0%, P < .001). In addition, pretreatment of tumor cells with the ligand CXCL12 enhanced the activation of the small GTPases Rho, Rac, and cdc42 as well as tumor cell extravasation without affecting tumor cell adhesion within liver sinusoids. Taken together, the findings of the present study provide first in vivo insights into the early events of chemokine ligand/receptor-mediated liver metastasis formation of tumor cells and define tumor cell extravasation rather than tumor cell arrest as the rate-limiting event.

In metastasis research, modern microscopic techniques shed a new light on the mechanisms of metastatic tumor cell arrest in the microcirculation of potential metastasis target organs. In this study, we differentiated the contribution of mechanical cell arrest, determined as lumen occlusion of liver sinusoids by tumor cells, and specific cell adhesion mediated by integrins for the arrest of human colon cancer cells in rat livers.

A safety margin of ≥10 mm is generally accepted in surgery for colorectal metastases. It is reasonable that modern methods of liver parenchyma dissection may allow for a reduction in this distance.

Metastatic lesions are the leading cause of death among cancer patients. These lesions usually originate from clonal proliferation of single tumor cells dispersed from the primary tumor into the circulation which finally arrest in the capillary bed of distant organs. The microenvironment within the circulation of potential metastatic target organs provides a variety of pro- and anti- metastatic stimuli regulating the onset of organ colonisation by metastatic tumor cells. Mechanical shear stress, anoikis and cell mediated cytotoxicity within the microcirculation probably clear most circulating tumor cells. Adhesion, and eventually extravasation, are essential initial interactions of circulating tumor cells with distant organs and can provide escape from the cytotoxic environment within the circulation. Adhesion to the capillary wall is mostly controlled by the organ-specific availability of adhesion molecules on tumor cells, the endothelium, and the composition of the underlying extracellular matrix. The availability of pro-adhesive and pro-migratory paracrine signals provided by the organ specific microenvironment can further initiate the onset of metastatic organ colonisation. Tumor cell and microenvironment factors regulating survival within the microcirculation, adhesion and extravasation of tumor cells are highlighted in the review.

Growing evidence supports substantial pathophysiological impact of platelets and their interactions on the development of septic lung failure. We developed a rat model of endotoxemia for direct in situ visualization of pulmonary microcirculation by in vivo fluorescence videomicroscopy. Male Sprague-Dawley rats were assigned to control, endotoxemia (Escherichia coli LPS, 15 mg/kg, i.v.), and fluid management for treatment of LPS-induced hypovolemia (Ringer lactate, hydroxyethyl starch [HES] 6%) groups (n = 7 each). Leukocytes were labeled in vivo by rhodamine, and 5 × 106 Calcein-AM-labeled nonactivated platelets were injected. Microcirculatory parameters (vessel diameter, ventilation-perfusion ratio) and adhesive characteristics of platelets and leukocytes (velocity, rolling, sticking) within the pulmonary microcirculation were quantified after endotoxin application under various regimens of fluid substitution for 60 min. A reduction of cell velocity and enhanced cell adhesion was seen in leukocytes and platelets (P < 0.05) after LPS injection. Fluid treatment with HES 6% resulted in a significant increase of platelet's velocity compared with the LPS group (442.86 ± 20.60 vs. 343.93 ± 11.17; P < 0.05), whereas Ringer lactate showed no beneficial effects. Similarly, HES 6% normalized LPS-induced platelet rolling and sticking as well as alterations in ventilation-perfusion ratio. Using direct visualization of the pulmonary microcirculation, we observed that platelet and leukocyte interactions are enhanced in the lung during LPS endotoxemia. Fluid therapy with HES 6% seems to have restorative effects on these cellular functions within the pulmonary microcirculation.

Abstract The endothelium is composed of a continuous, semipermeable, and metabolically highly active lining of endothelial cells (ECs) and an underlining basement membrane composed of various extracellular matrix components. The endothelium serves multiple physiologic functions in health and is fundamentally involved in many pathophysiologic conditions like inflammation; wound healing; and cancer, beside others. Endothelial cells mediate the nutritional supply of growing cancerous lesions, are involved in the body's immunologic response to cancer, and mediate initial interactions between circulating tumour cells and metastasis target organs. Endothelial cells and the basement membrane are not uniform throughout the body, but display distinct morphologic and functional characteristics depending on the vascular province they are derived from. Primary derived EC are usually not transformed and lose their endothelial differentiation during in vitro culture. Therefore, easily accessible macrovascular ECs like “human umbilical vein endothelial cells” (HUVEC) have been widely used in cancer research. These models are easily reproducible and standardized throughout the literature. Nevertheless, tumour cell–EC interactions are highly dependent on the ECs' origin and microvascular endothelial cells (MECs) show significant differences compared to macrovasular ECs in their morphologic and functional characteristics while interacting with other cell types like tumour cells. Therefore, while analysing tumour cell–EC interactions the ECs' source needs to be carefully selected according to the study's intention and has significant impact on experimental results obtained.

Introduction: Haematogenous homing of circulating tumor cells frequently accounts for intrahepatic recurrence of hepatocellular carcinoma after liver resection or transplantation for hepatocellular carcinoma. Understanding the molecular basis of this phenomenon is of crucial interest for the development of modern therapeutic strategies preventing tumor recurrence. Due to chemokine receptor expression, metastasing cancer cells can mimic leukocyte like behaviour like homing to different organs. In this study we evaluated the role of the chemokinereceptor CXCR4 for the homing of HCC cells to the liver in a rat model in vivo. Materials and methods: The hepatitis negative cell line HEP G2, derived from a human hepatocellular carcinoma was used. The chemokine receptor expression was determined by flowcytometry (FACS). Tumor cell adhesion and migration in vitro were determined in collagen and fibronectin coated static adhesion assays and trans well-chambers respectively. Tumor cell adhesion and migration in vivo were quantified by intravital fluorescence microscopy of the rat liver as described earlier. Localisation of the CXCR4 ligand CXCL12 in human livers was achieved by immunhistochemistry. After testing for normal distribution, mean values were compared by the T-test. Results: CXCR4 is expressed in 97 % of HEP G2 cells determined by FACS. The corresponding ligand CXCL12 exhibits chemotactic activity on HEP G2 cells seeded on collagen or fibronectin in a dose dependend manner at concentrations from 25 ng/ml to 100 ng/ml. Chemotaxis can be inhibited by anti-CXCR4 treatment of the cells. Adhesion of HEP G2 to collagen or fibronectin is not significantly influenced by anti-CXCR4 treatment. Immunhistochemical analysis of human livers without signs of cirrhosis or hepatitis showed significant CXCL12 expression in Kupffer cells and along the endothelial lining of the liver sinusoids. After intraarterial injection of fluorescene marked HEP G2 cells in rats, we observed adhesion of tumor cells within the hepatic sinusoids that was obviously not due to mechanical size restriction but specific adhesive interactions. When unspecific IgG control treated cells were used (n = 10), after 30 minutes 44,2 ± 3,4 cells/30 MF were arrested within the liver, while 22 % ± 4 % of these cells were extravadated into the liver parenchyma. After treatment of the cells with anti-CXCR4 IgG (n = 12) prior to injection, 49,4 ± 2,4 cells/30 MF were arrested in the liver after 30 minutes (n. s.), but the rate of extravadated cells was significantly reduced to 6 % ± 5 % (p < 0,05). Conclusions: Chemokine receptors, especially CXCR4, are expressed on human hepatocellular carcinoma. The only known Ligand for CXCR4, CXCL12, is expressed by sinusoidal endothelial cells and Kupffer cells and thereby perfedtly positioned within the hepatic microcirculation to support the homing of circulating tumor cells to the hepatic parenchyma.

Adhesive and invasive characteristics appear to be crucial for organ-specific metastasis formation. Using intravital microscopy we investigated the relation between the metastatic potential of colon carcinoma cells and their adhesive and invasive behavior during early steps of metastasis within microvasculatures of rat liver, lung, intestine, skin, muscle, spleen, and kidney in vivo. Colon carcinoma cells with low (HT-29P), intermediate (KM-12C), and high (HT-29LMM, KM-12L4) metastatic potential were injected into nude or Sprague-Dawley rats. Initial interactions with host organ microvasculatures were semiquantitatively analyzed throughout 20 to 30 minutes. Circulating cells passed microvessels in all observed organs without size restriction. All cell lines showed high adhesion rates, independent from their metastatic potential, within liver and lung but very rarely in other organs. Diameters of involved microvessels were larger than diameters of adherent tumor cells. Cell extravasation of highly metastatic HT-29LMM and KM-12L4 cells into liver parenchyma was significantly higher compared to low metastatic cells (P<0.05). Our results indicate that colon carcinoma cells can arrest in target organs without size restriction. Cell adhesion of circulating tumor cells occurred in metastatic target organs only, likely attributable to specific interactions. Migration into target organs correlated with their metastatic potential.

Growing evidence supports substantial pathophysiological impact of platelets (PLT) and their interactions on the development of septic lung failure. We developed a rat model of sepsis for direct in-situ visualization of pulmonary microcirculation by in vivo fluorescence videomicroscopy. Sprague-Dawley rats were assigned to control, sepsis (E. coli LPS, 15 mg/kg i.v.) and fluid management for circulatory stabilization (Ringer-Laktat, Haes 6%) groups (n = 8 each). Leukocytes were labeled in vivo by Rhoadamine (9 μmol/kg) and 5 × 107 Calcein-AM-labeled (1 μg/ml) non-activated PLT were injected. Microcirculatory parameters (eg. ventilation-perfusion-ratio) and characteristics of PLT and leukocytes (velocitity, adhaesion, rolling, sticking) within the pulmonary microcirculation were quantified after sepsis induction under various regimens of fluid substitution for 60 min. A reduction of cell velocity and enhanced cell adhaesion was seen in leukocytes and PLT (P < 0,05) after LPS injection. Fluid treatment with Haes 6% resulted in a significant increase of PLT velocity compared to the LPS group (442,86 ± 20,60 vs. 343,93 ± 11,17; P < 0,05) while Ringer-Laktat showed no beneficial effect. Similarly, Haes 6% normalized LPS-induced PLT rolling and sticking. Using direct visualization of the pulmonary microcirculation we observed that PLT and leukocyte interactions are enhanced during LPS-induced sepsis. Fluid therapy with Haes 6% seems to have restaurative effects on these cellular functions within the pulmonary microcirculation.

Introduction: Tumor cell migration and extravasation were recognized as fundamental cell properties involved in the formation cancer metastasis. A variety of different in vitro models has been established to monitor tumor cell motility and chemotaxis in 2- and 3-dimensional systems. Nevertheless in vitro systems are insufficient to analyze the complexity of the in vivo microenvironment. Using in vivo fluorescence video microscopy we therefore established an intravital model to semi-quantitatively analyze tumor cell migration within the rat liver. Materials and methods: Rat colon (CC531) and human colon carcinoma cell lines (HT-29 P, KM-12 C; their corresponding highly metastatic subclones: HT-29 LMM, KM-12 L4) were used. In male CD rats (250 g) a median laparotomy was performed and the left liver lobe placed under an in vivo fluorescence microscope without disturbance of the microcirculation. 106 fluorescence labeled (CalceinAM®) tumor cells were injected intraarterially. 30 standardized microscopic fields were analyzed in 5 min intervals for an observation period of 30 min. The migration rate was calculated as rate of migrated/extravasated cells in relation to all arrested cells. In some animals arrested cells were also detected by immunhistochemistry using a human specific anti-EPCAM antibody. Results: Using in vivo microscopy passing and arrested tumor cells within liver sinusoids were identified. Furthermore, arrested cells could be localized within the lumen of the sinusoids (»adherent cells«) or were seen extravasated (migrated cells) within the parenchyma. Immunhistochemistry confirmed the early extravasation of tumor cells. The rates of migrated cells increased over the 30 min observation period. Human and rat colon carcinoma cells demonstrated similar behavior in terms of cell adhesion and migration within the liver. Furthermore the high metastatic subclones demonstrated significantly higher migration rates than the parent cells: KM 12C (n = 7) 38% ± 7% vs. KM-12L4 (n = 4) 45% ± 5%, (p < 0.05) and HT-29 P (n = 9) 16% ± 11% vs. HT-29 LMM (n = 7) 29% ± 10%, (p < 0.05). Conclusions: Using this method metastatic tumor cell migration can be quantified in vivo. Cell lines with different metastatic potential demonstrated different migration rates suggesting tumor cell extravasation/migration as a rate limiting step of metastasis formation. This model makes it possible to study different factors that may be involved in metastatic tumor cell migration in vivo.

Tumor cells can show different malignant properties regarding their ability for organ-specific metastasis formation. Their adhesive and invasive characteristics mediated by various cell adhesion molecules appear to be crucial for this process. Using intravital fluorescence microscopy, we analyzed the adhesive and invasive interactions of circulating human colon carcinoma cells within the microvasculature of the liver in rats. The involvement of different cell adhesion molecules in specific tumor cell-host organ interactions was investigated. Single-cell suspensions of human colon carcinoma with low (HT-29P) and high (HT-29LMM) metastatic potential were fluorescence labeled with calcein-AM and intra-arterially injected into Sprague-Dawley rats. Initial interactions between different cell lines and the microvasculature of the liver were observed over 30 minutes and semiquantitatively analyzed. Different integrin subunits, carbohydrate ligands, and vascular cell adhesion molecule-1 were inhibited using function-blocking antibodies or by enzymatic removal. Inhibition of sialyl-Lewisa (sLea) or enzymatic removal of selectin carbohydrate ligands significantly reduced metastatic cell adhesion. In addition, α6-, β1-, and β4-integrins can directly mediate cell adhesion within the hepatic microcirculation. Furthermore,α2-,α6-,β1-, and β4-integrins are involved in early tumor cell extravasation into the liver parenchyma. Organ-specific formation of colorectal metastases appears to be mainly mediated by specific interactions between circulating carcinoma cells and the vessel wall of target organs but not mechanical entrapment. Selectins Lea interactions with sinusoidal endothelial cells can play a key role in organ-specific targeting, but direct integrin-mediated cell adhesion to extracellular matrix components in the space of Disse appears to be required for the successful formation of liver metastases.