Rat Liver Tumor Model Research Articles

Enhanced oral bioavailability and antitumor therapeutic efficacy of sorafenib administered in core-shell protein nanoparticle.

Orally delivered molecularly targeted small-molecule drugs play a significant role in managing cancer as a chronic disease. However, due to the poor oral bioavailability of some of these molecules, high-dose administration is required leading to dose-limiting toxicity especially when delivered daily for a long duration. Here, we report an oral nanoformulation for small-molecule multi-kinase inhibitor, sorafenib tosylate, showing nearly twofold enhancement in the oral bioavailability and enhanced therapeutic efficacy with a better safety profile compared to the current clinical formulation. Using a scalable process involving high-pressure homogenization, sorafenib was loaded into an albumin nanocarrier at ~ 50 w/w%. Repeated preparation of gram-scale batches (n = 7) showed an average particle size of 180 ± 9nm, encapsulation efficiency of 95 [Formula: see text] 2%, and drug-loading efficiency of 48 [Formula: see text] 0.7%. Further, surface engineering with a mucoadhesive layer on nanoparticles (referred to as ABSORF) resulted in the final size of 299 ± 38nm and surface charge of -54 ± 8mV. Single-dose and multidose pharmacokinetic studies showed twofold enhancement in the plasma concentration of sorafenib compared to current clinically used tablets. Antitumor efficacy studies in the orthotopic rat liver tumor model showed significant tumor regression (p value = 0.0037) even at half dose (eqv. to 200mg of human equivalent dose) of ABSORF compared to clinical control (eqv. to 400mg). The biodistribution of sorafenib from ABSORF was higher in the liver; however, liver and kidney function test parameters were comparable with that of the 2 × dose of clinical control. No abnormalities and signs of toxicity were seen in the histopathological analysis for ABSORF-treated animals. In summary, we demonstrate a scalable preparation of small-molecule drug-loaded nanoformulation with approximately twofold enhancement in oral bioavailability, improved antitumor efficacy, and acceptable toxicity profile.

Safety and Feasibility of Contrast-Enhanced Computed Tomography with a Nanoparticle Contrast Agent for Evaluation of Diethylnitrosamine-Induced Liver Tumors in a Rat Model

Safety and feasibility of contrast-enhanced computed tomography (CECT) with a nanoparticulate contrast agent, ExiTron nano 12000, was evaluated in a rat liver tumor model. This study employed eighteen 8-week-old male F344 rats. Six rats given tap water for 8 weeks further divided into two: Control group and Normal Liver with CECT group. Six rats each were given tap water containing diethylnitrosamine (DEN) at 100 ppm for 8 or 14 weeks; Adenoma group and Hepatocellular carcinoma (HCC) group, respectively. Biochemical marker values and adverse events were evaluated after CT imaging. ExiTron nano 12000 was evaluated for the hepatic contrast enhancement, and the detection and measurement of liver nodules by CECT after 8- and 14-weeks administration of DEN. Post-mortem liver specimens were evaluated by hematoxylin-eosin (HE) staining, and the number and size of liver nodules were measured. The HCC group was evaluated for diagnostic concordance between HE-stained and CECT-detected nodules. The contrast agent enhanced liver and was tolerated after CECT in 15 rats. Biochemical parameter values did not differ significantly between the Control and Normal Liver groups. The numbers of CECT-detected nodules in the Adenoma and HCC groups were 14.8 ± 5.1, and 32.4 ± 8.1, respectively. The HCC group had 3.6 ± 2.7 of pathological HCCs, which were identified by CECT. The size of CECT-detected HCCs correlated significantly with that of pathological HCCs (r = 0.966, p < 0.0001). CECT with ExiTron nano 12000 is a safe and feasible method to measure tumors in a rat liver tumor model.

Abstract No. 266 MRI combined with PET for prediction of outcomes after irreversible electroporation in a rat liver tumor model

Abstract No. 266 MRI combined with PET for prediction of outcomes after irreversible electroporation in a rat liver tumor model

Ultrasonic Heating Detects Lipiodol Deposition within Liver Tumors after Transarterial Embolization: An In Vivo Approach.

Simple SummaryThe accumulation of Lipiodol (ethiodized oil) after transarterial embolization is known to reflect tumor necrosis. In general, the treatment effect is evaluated by computed tomography; there has been no development in imaging modalities for several decades. A new technique, ultrasonic heating, can differentiate biological tissues based on the fact that tissues’ characteristic sound velocity varies depending on the temperature. This technique could have the potential to evaluate treatment effect after transarterial embolization as an alternative to computed tomography.Computed tomography (CT) is the standard method to evaluate Lipiodol deposition after transarterial embolization (TAE) for a long period. However, iodine but not Lipiodol can be observed on CT. A minimally invasive other method to detect Lipiodol has been needed to evaluate accurate evaluation after procedure. The purpose of this study was to evaluate the efficacy of using the rate of change in sound velocity caused by ultrasonic heating to reflect Lipiodol accumulation after TAE in a rat liver tumor model. We analyzed the association of this developed technique with CT images and histological findings. Eight rats bearing N1S1 cells were prepared. After confirmation of tumor development in a rat liver, Lipiodol was injected via the hepatic artery. Seven days after TAE, CT scan and sound velocity changes caused by ultrasonic heating were measured, and then the rats were sacrificed. An ultrasonic pulse-echo method was used to measure the sound velocity. The temperature coefficient of the sound velocity in each treated tumor was evaluated and compared with the mean CT value and the histological Lipiodol accumulation ratio. Pearson’s correlation coefficients were calculated to assess the correlation between the measured values. The correlation coefficient (r) of the mean CT value and histological Lipiodol accumulation ratio was 0.835 (p = 0.010), which was considered statistically significant. Also, those of the temperature coefficient of the sound velocity and the histological Lipiodol accumulation ratio were statistically significant (r = 0.804; p = 0.016). To our knowledge, this is the first study that reported the efficacy of ultrasonic heating to detect Lipiodol accumulation in rat liver tumors after TAE. Our results suggest that the rate of change in sound velocity caused by ultrasonic heating can be used to evaluate Lipiodol accumulation in liver tumors after TAE, and thus could represent an alternative to CT in this application. This new innovative technique is easy to treat and less invasive in terms of avoiding radiation compared with CT.

Useful Parameters in Dynamic Contrast-enhanced Ultrasonography for Identifying Early Response to Chemotherapy in a Rat Liver Tumor Model.

Objectives:The objective of the study is to determine a parameter on the time-intensity curve (TIC) of dynamic contrast-enhanced ultrasonography (DCE-US) that best correlates with tumor growth and to evaluate whether the parameter could correlate with the early response to irinotecan in a rat liver tumor model.Material and Methods:Twenty rats with tumors were evaluated (control: Saline, n = 6; treatment: Irinotecan, n = 14) regarding four parameters from TIC: Peak intensity (PI), k value, slope (PI × k), and time to peak (TTP). Relative changes in maximum tumor diameter between day 0 and 10, and parameters in the first 3 days were evaluated. The Mann-Whitney U-test was used to compare differences in tumor size and other parameters. Pearson’s correlation coefficients (r) between tumor size and parameters in the control group were calculated. In the treatment group, relative changes of parameters in the first 3 days were compared between responder and non-responder (<20% and ≥20% increase in size on day 10, respectively).Results:PI, k value, PI × k, and TTP significantly correlated with tumor growth (r = 0.513, 0.911, 0.665, and 0.741, respectively). The mean RC in k value among responders (n = 6) was significantly lower than non-responders (n = 8) (mean k value, 4.96 vs. 72.5; P = 0.003).Conclusion:Parameters of DCE-US could be a useful parameter for identifying early response to irinotecan.

03:09 PM Abstract No. 422 Early prediction of tumor response using contrast-enhanced ultrasonography in intra-arterial therapy by micellar nanoparticles in a rat liver tumor model

To determine the best parameter to collate changes of tumor sizes in time intensity curve (TIC) of contrast-enhanced ultrasonography (CEUS) and to evaluate whether the determined parameter could predict tumor response in intra-arterial micellar nanoparticles incorporating SN-38 in a rat colorectal liver metastasis model. Eleven rats with colorectal liver metastasis were evaluated (control group: intravenous saline, n=4, and treatment group; intra-arterial micellar nanoparticles via hepatic artery, n=7). The maximum diameter of each tumor was measured on pre, 3 and 10 days after treatment. In the control group, a correlation coefficient (r) was calculated in the following 4 parameters measured on day 3; the peak intensity (PI), k value, the slope of wash-in (PI x k) and time to peak (T to Pk). The relationship between the changes of tumor diameter from pre to 10 days and the parameters were evaluated, then the most sensitive parameter was determined. In the treatment group, the best parameter obtained on day 3 was compared between the responder group (less than 20% decreased tumor size on day 10) and the non-responder group (more than 20% increased tumor size). In the control group, all tumors increased. The increased ratios were +2.25% on day 3 and +61.6% on day10. The correlation coefficients of parameters were 0.144 in PI, 0.870 in k, 0.709 in PI x k and -0.363 in T to Pk. The k value was determined as the most sensitive parameter. In the treatment group, the mean tumor increased ratios on day 3 and day 10 in the responder group (n=4) were -0.17% and -15.1%, respectively, while those of in the non-responder group (n=3) were +1.32% and +23.2%, respectively. The mean k value in the responder group on day 3 was significantly lower than that in the non-responder group (mean value; -0.057 vs 0.422, p=0.034). The k value in TIC obtained by CEUS can be a sensitive parameter to predict tumor response in intra-arterial therapy using micellar nanoparticles in rat model. Further clinical investigation is mandatory.Tabled 1Correlation Coefficient Between the Relative Change of Size and of the Parameters in TICSize (average)RC % of Size (average)RC of the Parameters (3 days, average)Day 0Day 3Day 103 days10 daysT to PkkPISlopeControl11.0511.317.852.2561.6-0.3883.250.2023.575Correlation coefficientr-0.3630.870.1440.709Size (average)RC % of Size (average)RC of the Parameters (3 days, average)Day 0DayDay 103 days10 daysT to PkkPISlopeResponder11119.625-0.171-15.10.0989-0.05710.302-0.117Non-responder10.110.212.41.3223.20.01120.4220.3990.597Correlation coefficientr-0.7950.7150.470.461RC; relative changeT to Pk; time to peak, PI; peak intensity, slope = k x PI Open table in a new tab

Higher CYP2E1 Activity Correlates with Hepatocarcinogenesis Induced by Diethylnitrosamine.

Hepatocellular carcinoma (HCC) is one of the most common types of primary liver cancer and the third most frequent cause of cancer death worldwide. Diethylnitrosamine (DEN) is one of the recognized risk factors for hepatocarcinogenesis likely due to CYP2E1-mediated metabolic activation. However, CYP2E1-mediated DEN metabolic activity in non-neoplastic liver tissue from HCC patients has not been determined; the role of CYP2E1 activity, in particular CYP2E1 constitutive activity and CYP2E1 inhibited activity, with respect to the hepatocarcinogenesis induced by DEN is not yet clear. Herein, we determined CYP2E1-mediated DEN metabolic activity in non-neoplastic liver tissue from HCC patients and found that CYP2E1-mediated DEN metabolic activity was significantly elevated with a 43.3% positive rate, and clinicopathologic parameters did not affect the activity. Then, using a Sprague-Dawley rat liver tumor model induced by DEN, the relationship between CYP2E1 constitutive/inhibited activity and hepatocarcinogenesis was explored. The results showed that the CYP2E1 constitutive activity was strongly correlated with tumor incidence and severity of liver tumorigenesis (nodule numbers and size), whereas inhibition of CYP2E1 activity decreased hepatocyte proliferation, liver injury, and liver carcinogenesis in the presence of DEN. In conclusion, the higher CYP2E1 activity would lead to an increased incidence of HCC as a result of CYP2E1-mediated DEN activation. Therefore, higher CYP2E1 activity might be a risk factor for HCC induced by DEN.

Abstract No. 670 - Transarterial chemoembolization with parthenolide in a rat liver tumor model induces tumor regression without any detectable liver or systemic toxicity

Abstract No. 670 - Transarterial chemoembolization with parthenolide in a rat liver tumor model induces tumor regression without any detectable liver or systemic toxicity

Experimental Study Using Rat Liver Tumor Model for Interventional Radiologists

Experimental Study Using Rat Liver Tumor Model for Interventional Radiologists

Low-dose splenic radiation inhibits liver tumor development of rats through functional changes in CD4+CD25+Treg cells

The increased number of CD4+CD25+Treg cells in tumor local and peripheral splenic tissues is related to the low immune function as well as to tumor recurrence and metastasis. Our pre-clinical studies showed that low-dose radiation (LDR) of the spleen in liver cancer patients significantly improves immune functions. However, the molecular mechanisms of such radiation remained ill defined. This study explores the role of CD4+CD25+Treg cells in radiation-induced immunomodulatory effects. Using the diethylnitrosamine (DEN)-induced rat liver tumor model and in vitro cell experiments, the percentage of CD4+CD25+Treg/CD4+ cells in the blood and the expressions of Foxp3+, IL-10, TGF-β, and cytotoxic T lymphocyte-associated antigen-4(CTLA-4) in spleen and liver tumors significantly decreased after LDR of the spleen in rats with liver cancer. The tumors became smaller than those in the non-radiated group, with both showing a parallel relation. Flow cytometry and MTT results revealed that LDR failed to inhibit CD4+CD25+Treg cell proliferation. Conversely, apoptosis was reduced and proliferation was stimulated. This process also changed CTLA-4 molecule expression on the surfaces of CD4+CD25+Treg cells and reduced their inhibitory function against CD4+CD25−T cell proliferation, and the suppression function of CD4+CD25+Treg cells was further weakened with the introduction of the CTLA-4 inhibitor. Findings demonstrate that the reduction of CTLA-4 expression on the CD4+CD25+Treg cell surface and the further inhibition of cell function may be considered as important regulators of LDR-induced immunomodulatory effects. This study provides experimental evidence to elucidate the immune enhancement induced by this process and presents a novel method for liver cancer immunotherapy.

Hemodynamics in rat liver tumor model during retrograde-outflow isolated hepatic perfusion with aspiration from the portal vein: angiography and in vivo microscopy

Orthograde percutaneous isolated hepatic perfusion (IHP) techniques using balloon occlusion catheters are relatively simple and facilitate repeated therapy, but they result in higher rates of leakage from the perfusion circuit into the systemic circulation. Therefore, a feasible protocol for percutaneous IHP with less leakage is required. To investigate hemodynamic changes in rat liver and tumor during retrograde-outflow isolated hepatic perfusion (R-IHP) with aspiration from the portal vein (PV). Animal experiments were approved by the Animal Experiment Ethics Committee of Lund University. Eighteen rats underwent R-IHP after laparotomy and catheterization of the PV and hepatic artery (HA). The HA, inferior vena cava (IVC), and PV were ligated, and flow through the suprahepatic IVC was controlled with a suture loop. The rats were divided into two groups to examine blood flow during R-IHP. Four rats (group 1) underwent arteriography via the HA with and without R-IHP, and 14 rats (group 2) were inoculated with tumor and examined by in vivo fluorescence microscopy of liver and tumor during R-IHP. In group 1, hepatic arteriography during R-IHP confirmed arterioportal communication in all four rats, with the PV acting as an outflow tract. In vivo fluorescence microscopy in group 2 showed strong enhancement of tumors, and no blood supply from the portal venules to the tumors was seen in any of the 14 rats. Blood flow in the major portion of the hepatic lobules was stopped and the percentage of enhanced area was significantly lower in the normal hepatic lobules than in the tumors (P < 0.0001). We confirmed reversal of blood flow concomitant with good perfusion of the liver tumor and with reduced perfusion of normal liver parenchyma during R-IHP.

A novel oxygen carrier "YQ23" suppresses the liver tumor metastasis by decreasing circulating endothelial progenitor cells and regulatory T cells.

BackgroundSurgical therapies are the first-line treatments for hepatocellular carcinoma (HCC) patients. However, the high incidence of tumor metastasis after liver surgery remains a severe problem. We aim to investigate the roles and the underlying mechanism of YQ23, stabilized non-polymeric diaspirin cross-linked tetrameric hemoglobin, in liver tumor metastasis after major hepatectomy and partial hepatic ischemia reperfusion (I/R) injury.MethodsAn orthotopic liver tumor model in Buffalo rat was established using the hepatocellular carcinoma cell line McA-RH7777. Major hepatectomy for tumor-bearing lobe and partial hepatic I/R injury were performed at two weeks after orthotopic liver tumor implantation. YQ23 (0.2 g/kg) was administered at 1 hour before ischemia and immediately after reperfusion. Blood samples were collected at day 0, 1, 7, 14, 21 and 28 for detection of circulating endothelial progenitor cells (EPCs) and regulatory T cells (Tregs).ResultsOur results showed that YQ23 treatment effectively inhibited intrahepatic and lung metastases together with less tumor angiogenesis at 4 weeks after major hepatectomy and partial hepatic I/R injury. The levels of circulating EPCs and Tregs were significantly decreased in YQ23 treatment group. Furthermore, YQ23 treatment also increased liver tissue oxygenation during hepatic I/R injury. Up-regulation of HO1 and down-regulation of CXCR3, TNF-α and IL6 were detected after YQ23 treatment.ConclusionsYQ23 treatment suppressed liver tumor metastasis after major hepatectomy and partial hepatic I/R injury in a rat liver tumor model through increasing liver oxygen and reducing the populations of circulating EPCs and Tregs.

Different roles of hepatic hypothermic ischemia and ischemic preconditioning in chemically induced hepatocarcinogenesis in rats

BackgroundHepatic ischemia–reperfusion (IR) injury, an unfavorable complication of hepatectomy, could be prevented by hypothermic ischemia and ischemic preconditioning (IPC). However, the effects of these two approaches on hepatocarcinogenesis have not been examined. The aim of the study was to investigate roles of hypothermic ischemia and IPC in a chemically induced rat liver tumor model. MethodsTwenty-four Sprague–Dawley rats were treated with diethylnitrosamine and phenobarbital to induce hepatocellular carcinoma. Rats underwent hepatic ischemic injury, hypothermic ischemia, and IPC. Twenty-eight-wk-old rats were sacrificed to evaluate the morbidity and growth of liver tumor. Cytokines were measured at the protein and messenger RNA level. ResultsIR injury significantly promoted liver tumor development. Intriguingly, hypothermic ischemia, but not IPC, delayed liver carcinogenesis, although both of them suppressed the hepatic IR injury. IPC-treated rats showed elevated interleukin (IL)-6 concentration in the serum and messenger RNA expression in liver. In addition, higher levels of IL-6 activated signal transducer and activator of transcription 3 in the liver of IPC-treated rats. The hepatic expression of target genes of signal transducer and activator of transcription 3 signaling, cyclin D1, c-myc, c-fos, and c-jun, all of which might participate in tumor progression, increased in IPC group, compared with that of IR group. ConclusionsThese data indicated hypothermic ischemia could ameliorate both IR injury and liver tumor development. However, IPC, another effective method to prevent hepatic IR injury, might exacerbate liver tumor growth. The elevated level of IL-6 was one of the reasons for the different effects of hypothermic ischemia and IPC on hepatocarcinogenesis in rats.

Multimodality Imaging to Assess Immediate Response to Irreversible Electroporation in a Rat Liver Tumor Model

To compare changes on ultrasonographic (US), computed tomographic (CT), and magnetic resonance (MR) images after irreversible electroporation (IRE) ablation of liver and tumor tissues in a rodent hepatoma model. Studies received approval from the institutional animal care and use committee. Forty-eight rats were used, and N1-S1 tumors were implanted in 24. Rats were divided into groups and allocated for studies with each modality. Imaging was performed in normal liver tissues and tumors before and after IRE. MR imaging was performed in one group before and after IRE after hepatic vessel ligation. US images were graded to determine echogenicity changes, CT attenuation was measured (in Hounsfield units), and MR imaging signal-to-noise ratio (SNR) was measured before and after IRE. Student t test was used to compare attenuation and SNR measurements before and after IRE (P < .05 indicated a significant difference). IRE ablation produced greater alterations to echogenicity in normal tissues than in tumors. Attenuation in ablated liver tissues was reduced compared with that in control tissues (P < .001), while small attenuation differences between ablated (42.11 HU ± 2.11) and control (45.14 HU ± 2.64) tumors trended toward significance (P = .052). SNR in ablated normal tissues was significantly altered after IRE (T1-weighted images: pre-IRE, 145.95 ± 24.32; post-IRE, 97.80 ± 18.03; P = .004; T2-weighted images, pre-IRE, 47.37 ± 18.31; post-IRE, 90.88 ± 37.15; P = .023). In tumors, SNR differences before and after IRE were not significant. No post-IRE signal changes were observed after hepatic vessel ligation. IRE induces rapid changes on gray-scale US, unenhanced CT, and MR images. These changes are readily visible and may assist a performing physician to delineate ablation zones from the unablated surrounding parenchyma.

Transcatheter arterial embolization promotes liver tumor metastasis by increasing the population of circulating tumor cells

Transcatheter arterial embolization (TAE) is widely used as an effective palliative treatment for hepatocellular carcinoma (HCC), and can prolong survival time. However, the high incidence of tumor recurrence and metastasis after TAE is still a major problem. Recent studies demonstrated that circulating tumor cells (CTCs) contribute to tumor metastasis. In this study, we tried to clarify whether the residual HCC after TAE can increase metastasis by increasing the number of CTCs. An orthotopic liver tumor model in the Buffalo rat was established using green fluorescent protein (GFP)-transfected HCC cell line, McA-RH7777. Two weeks after orthotopic liver tumor implantation, the rats underwent TAE treatment from the gastroduodenal artery. Iodized oil or saline was injected intra-arterially. Blood samples were taken on day 0, 1, 3, 7, 14, and 21 for detection of CTCs after TAE treatment. We analyzed the number of CTCs and assessed the metastatic potential of surviving tumor cells in rats between TAE and control groups. Our results demonstrated that the metastatic colonies in the lung were significantly increased by TAE treatment. The number of CTCs was also significantly increased by TAE treatment from day 7 to day 21. The expression of hypoxia-inducible factor (HIF)-1α and epithelial–mesenchymal transition (EMT) marker proteins (N-cadherin and vimentin) was upregulated, but E-cadherin was downregulated after TAE treatment. In conclusion, the metastatic potential of residual HCC can be induced by TAE treatment in a rat liver tumor model, which involves the acquisition of EMT features and an increased number of CTCs.

Abstract No. 270 - Multimodality imaging to assess immediate response to irreversible electroporation in a rat liver tumor model

Purpose The purpose was to compare induced changes within US, CT, and MRI images shortly after irreversible electroporation (IRE) ablation of normal liver parenchyma and hepatic tumors. Materials and Methods All procedures were approved by IACUC. 48 rats were divided into 8 groups, 24 were implanted with N1-S1 tumors. US, CT, and MRI measurements were performed in both normal and tumor tissues pre/post-IRE. MRI was also performed in one group pre/post-IRE after hepatic vessel ligation. Pre/post-IRE US images were graded by 3 radiologists to determine echogenicity changes (0: no change; 1: slight change; 2: clear change; 3: dramatic change). Hounsfield units (HU) were measured based on CT and signal to noise ratio (SNR) measured within T2W MR images pre/post-IRE. Results The average grade was 2.0 for echogenicity change in ablated normal liver and 1.3 for ablated tumor. HU of ablated normal liver (62.53±2.24) was significantly different (p=0.0001) from unablated parenchyma (46.99±5.259); while small, HU difference between ablated tumor (42.11±2.11) and control tumor (45.14±2.64) did trend toward significance (p=0.0524). SNR in ablated normal liver was significantly increased post-IRE (PRE: 47.37±18.31, POST: 90.88±37.15, p=0.0227). SNR difference between tumors pre/post-IRE was not significant (PRE: 102.13±31.36, POST: 101.09±21.66, p=0.9401). No post-IRE signal changes were observed for rats that underwent portal vein and hepatic artery ligation prior to MRI-monitored IRE. IRE ablation zones were confirmed with HandE and micro-structural changes observed with transmission electron microscopy. Conclusion IRE induces rapid changes within US, CT, and MRI images. However, these imaging methods characterized IRE-induced signal changes more effectively in normal parenchyma than in tumors. MRI findings following hepatic ligation indicate that early signal changes are likely the result of local fluid accumulation in the treated zones. Future studies are necessary to develop superior imaging methods to better characterize electroporation-induced changes in tumors.

Abstract No. 266 - Evaluation of the microcirculation and perfusion of a rat liver tumor model after transarterial embolization using contrast-enhanced ultrasonography and perfusion analysis

Purpose To evaluate the feasibility of contrast enhanced ultrasound in evaluation the changes of microcirculation and perfusion after transarterial embolization (TAE) in an N1-S1 model of hepatocellular carcinoma in rats. Materials and Methods This study was approved by the Institutional Animal Care and Use Committee at Case Western Reserve University. N1-S1 cells (5 × 10^6) were inoculated under the capsule of the median aspect of the left liver lobe of 10 Sprague-Dawley rats (6 weeks old, Charles River, Wilmington, MA) after a laparotomy. Two weeks later, the tumors were embolized by arterial injection of PVA particles (45-150 μm, Contour, Boston Scientific) after retrograde catheterization via gastroduodenal artery. Contrast enhanced ultrasound (CEUS) was performed before and 3 days after TAE. A prototype perfusion software (ImageLab, Toshiba Medical Systems Corp.) was used to analyze the first-pass enhancing images. Time-signal intensity-curves in the tumor and normal liver were first obtained, and then fitted with an S-shape function. The perfusion parameters [peak, mean transit time (MTT), flow] pre and 3 days after TAE were then calculated. Results Seven tumors (9.3 - 20.1 mm in maximum diameter) were established in the left lobes of rats. The other three animals without tumors were excluded from the study. On CEUS imaging, all liver tumors were enhanced before TAE. A significant filling defect and reduced blood flow in the whole tumor was noted on day 3 after TAE. The changes of parameters in normal liver and the MTT in tumor after TAE were not significant. The time to peak and flow in the tumors was significantly decreased after TAE (1.39 ± 1.05 vs 0.01 ± 0.03, 0.45 ± 0.40 vs 0.00 ± 0.01, respectively). The normalized tumor flow (flow in tumor / flow in liver) was 1.95 ± 1.71 vs 0.05 ± 0.08 pre and post TAE (P = 0.01), and the change rate was -96.47% ± 7.65. Conclusion CEUS and quantitative perfusion analysis is a useful and convenient tool to evaluate the microcirculation change after TAE, especially in hypervascular liver lesions.

Enhanced Antitumor Efficacy of a Vascular Disrupting Agent Combined with an Antiangiogenic in a Rat Liver Tumor Model Evaluated by Multiparametric MRI

A key problem in solid tumor therapy is tumor regrowth from a residual viable rim after treatment with a vascular disrupting agent (VDA). As a potential solution, we studied a combined treatment of a VDA and antiangiogenic. This study was approved by the institutional ethical committee for the use and care of laboratory animals. Rats with implanted liver tumors were randomized into four treatment groups: 1) Zd6126 (Zd); 2) Thalidomide (Tha); 3) Zd in combination with Tha (ZdTha); and 4) controls. Multiparametric MRIs were performed and quantified before and after treatment. Circulating endothelial progenitor cells (EPCs) and plasma stromal cell-derived factor-1α (SDF-1α) were monitored. Tumor apoptosis, necrosis, and microvessels were verified by histopathology. A single use of Zd or Tha did not significantly delay tumor growth. The combined ZdTha showed enhanced antitumor efficacy due to synergistic effects; it induced a cumulative tumor apoptosis or necrosis, which resulted in significant delay in tumor growth and reduction in the viable tumor rim; it also reduced tumor vessel permeability; and it improved tumor hemodynamic indexes, most likely via a transient normalization of tumor vasculature induced by Tha. A stepwise linear regression analysis showed that the apparent diffusion coefficient was an independent predictor of tumor growth. We found no significant increases in Zd-induced circulating EPCs or plasma SDF-1α. ZdTha showed improved therapeutic efficacy in solid tumors compared to either agent alone. The therapeutic effects were successfully tracked in vivo with multiparametric MRI.

FTY720 Suppresses Liver Tumor Metastasis by Reducing the Population of Circulating Endothelial Progenitor Cells

BackgroundSurgical procedures such as liver resection and liver transplantation are the first-line treatments for hepatocellular carcinoma (HCC) patients. However, the high incidence of tumor recurrence and metastasis after liver surgery remains a major problem. Recent studies have shown that hepatic ischemia-reperfusion (I/R) injury and endothelial progenitor cells (EPCs) contribute to tumor growth and metastasis. We aim to investigate the mechanism of FTY720, which was originally applied as an immunomodulator, on suppression of liver tumor metastasis after liver resection and partial hepatic I/R injury.Methodology/Principal FindingsAn orthotopic liver tumor model in Buffalo rat was established using the hepatocellular carcinoma cell line McA-RH7777. Two weeks after orthotopic liver tumor implantation, the rats underwent liver resection for tumor-bearing lobe and partial hepatic I/R injury. FTY720 (2 mg/kg) was administered through the inferior caval vein before and after I/R injury. Blood samples were taken at days 0, 1, 3, 7, 14, 21 and 28 for detection of circulating EPCs (CD133+CD34+). Our results showed that intrahepatic and lung metastases were significantly inhibited together with less tumor angiogenesis by FTY720 treatment. The number of circulating EPCs was also significantly decreased by FTY720 treatment from day 7 to day 28. Hepatic gene expressions of CXCL10, VEGF, CXCR3, CXCR4 induced by hepatic I/R injury were down-regulated in the treatment group.Conclusions/SignificanceFTY720 suppressed liver tumor metastasis after liver resection marred by hepatic I/R injury in a rat liver tumor model by attenuating hepatic I/R injury and reducing circulating EPCs.

Free-Breathing Quantitative Dynamic Contrast-Enhanced Magnetic Resonance Imaging in a Rat Liver Tumor Model Using Dynamic Radial T1 Mapping

: The high sensitivity to motion artifacts is a major limiting factor for applying the dynamic 3D T1-weighted gradient-echo (3D T1w GRE) technique for dynamic contrast-enhanced (DCE) magnetic resonance imaging (MRI) experiments in small rodents. Dynamic quantification of the relaxation rate R1 (1/T1) presents an alternative approach to reduce these motion artifacts. In this work, an optimized 2D single-shot Look-Locker based T1 mapping technique, named GOLD, applying radial sampling in the golden-angle view order and contrast-enhancing k-space filter was evaluated for its use in free-breathing quantitative DCE-MRI of rat liver on a clinical 1.5 T MRI system. : In vitro measurements and initial in vivo experiments in healthy rats were performed to evaluate the accuracy and resilience of the GOLD technique to motion artifacts. Unifocal hepatocellular carcinoma (HCC) was established in 20 male Buffalo rats. Twelve days after tumor cell implantation, animals were screened for intrahepatic tumor nodules by high-resolution T2-weighted MRI. Quantitative DCE-MRI experiments applying bolus injected gadopentetate dimeglumine were performed in 11 HCC-bearing rats using the GOLD technique. For comparison, a standard 3D T1w GRE sequence was applied in 6 additional rats. : Phantom experiments showed good agreement for T1 values measured by the GOLD method and an inversion recovery spectroscopy measurement. The in vivo experiments in healthy rats confirmed the robustness of the GOLD method in T1 value determination and its resilience to motion artifacts. Gadopentetate dimeglumine concentration (CGd) time curves determined from free-breathing GOLD-based DCE-MRI experiments of HCC-bearing rats allowed reliable and robust pharmacokinetic modeling (K, ve, lag time Td, and slow washout rate rwo) of tumor, liver, and spinal muscle. In comparison to the dynamic 3D T1w GRE, the GOLD method showed less variation and jitter in the CGd time curves and significantly increased accuracy (in terms of the goodness of fit) in the pharmacokinetic modeling. Significant differences were detected for K and ve with the 3D T1w GRE method apparently underestimating those parameters. : The GOLD technique allowed dynamic sampling of 2D axial T1 maps of the rat abdomen with 6-second temporal resolution enabling simultaneous and robust pharmacokinetic modeling of HCC, normal liver, and spinal muscle.