Role of microRNAs in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer related death worldwide. HCC develops through a multistep process that involves genetic and epigenetic changes. In addition to genetic and epigenetic mechanisms, recent studies have shown that microRNAs (miRNAs) play essential roles in hepatocellular carcinogenesis through the post-transcriptional regulation of tumor associated-genes. In this review, we summarize the role of miRNAs in HCC and its microenvironment, and discuss the implications for HCC therapy.

Differential diagnosis of gallbladder wall thickening: the usefulness of contrast-enhanced ultrasound.

he purpose of this study was to evaluate the usefulness of contrast-enhanced ultrasound (CEUS) in the differential diagnosis of gallbladder wall (GBW) thickening and determine the predictors of malignant GBW thickening. One hundred fifty-nine patients with GBW thickening, including 76 men and 83 women, from eight institutions were enrolled. CEUS was performed after injection of a sulfur hexafluoride microbubble-based ultrasound contrast agent. Multiple logistic regression analysis was used to reveal independent predictor sassociated with malignant GBW thickening. The final diagnoses were 48 gallbladder carcinomas and 111 benign gallbladder diseases.Maximal thicknesses of the GBW in malignant and benign GB Wthickening were 17.3 ± 5.2 (6 ­ 30) mm and 8.6 ± 5.1 (4 ­ 26) mm respectively (p , 0.001). CEUS revealed significant differences in intralesional vessels, enhancement homogeneity, time to hypo-enhancement, inner layer discontinuity, outer layer discontinuity and adjacent liver involvement (all p-values , 0.05) between malignant and benign GBW thickening. Patient age . 46.5 y, focal GBW thickening, inner layer discontinuity and outer layer discontinuity were found to be associated with malignancy by multiple logistic regression analysis (all p-values , 0.05). Receiver operating characteristic curve analysis revealed Az values for patient age, focal GBW thickening, inner wall discontinuity and outer wall discontinuity of 0.709 (95%confidence interval [CI]: 0.627­0.790), 0.714 (95% CI: 0.630­0.798), 0.860 (95%CI: 0.791 ­ 0.928) and 0.858 (95% CI: 0.783 ­ 0.933), respectively. CEUS is useful in the differential diagnosis between malignant and benign GBW thickening. Focal GBW thickening, inner wall discontinuity and outer wall discontinuity observed on CEUS are diagnostic clues for malignant GBW thickening.

Bipolar radiofrequency ablation for liver tumors: comparison of contrast-enhanced ultrasound with contrast-enhanced MRI/CT in the posttreatment imaging evaluation.

OBJECTIVE: The aim of the study was to assess the role of contrast-enhanced ultrasound (CEUS) in treatment response evaluation after percutaneous bipolar radiofrequency ablation (BRFA) for liver tumors. METHODS: From May 2012 to May 2014, 39 patients with 73 tumors were treated by BRFA. One month after the treatment, CEUS and CEMRI/CECT were conducted to evaluate the treatment response. The results of CEUS were compared with CEMRI/CECT. RESULTS: Of the 73 tumors ablated, eight (11.0%) were found to have residual viable tumor tissue and 65 (89.0%) were successfully ablated based on CEMRI/CECT within 1-month after ablation. CEUS detected seven of the eight residual tumors and 63 of 65 completely ablated tumors. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy of CEUS were 87.5% (7/8), 96.9% (63/65), 77.8% (7/9), 98.4% (63/64) and 95.9% (70/73), respectively. The complete ablation (CR) rates for the tumors ≤ 3.0 cm, 3.1-5.0 cm, and >5.0 cm were 96.6% (58/60), 63.6% (7/11), and 0% (0/2), respectively (P<0.001). CR rates were 94.7% (36/38) for primary liver tumors and 82.9% (29/35) for metastatic liver tumors (P=0.212), and were 97.4% (38/39) for the tumors with curative treatment intention and 79.4% (27/34) for those with palliative treatment intention (P=0.037). Major complication was not encountered in this series. CONCLUSIONS: BRFA is an effective technique of percutaneous ablation for liver tumors and CEUS can be used to assess its therapeutic effect accurately.

Acoustic radiation force impulse imaging: a new tool for the diagnosis of papillary thyroid microcarcinoma.

PURPOSE: To evaluate the diagnostic performance of ARFI imaging in differentiating between benign and malignant thyroid nodules <1 cm. MATERIALS AND METHODS: 173 pathologically proven thyroid nodules (77 benign, 96 malignant) in 157 patients were included in this study. Receiver-operating characteristic curve (ROC) analyses were performed to assess the diagnostic performance of conventional ultrasound (US) and ARFI imaging in papillary thyroid microcarcinoma (PTMC). The independent risk factors for predicting PTMC were evaluated. RESULTS: The mean SWV value of benign and malignant thyroid nodules were 2.57 ± 0.79 m/s (range: 0.90-4.92 m/s) and 3.88 ± 2.24 m/s (range: 1.49-9.00 m/s) (P = 0.000). Az for VTI elastography score was higher than that for hypoechoic, absence of halo sign, and type III vascularity (P < 0.05). The optimal cut-offs for VTI elastography score and SWV were score 4 and 3.10 m/s. Gender, hypoechoic, taller than wide, VTI elastography score ≥ 4, and SWV > 3.10 m/s had been found to be independent risk factors for predicting PTMC. CONCLUSION: ARFI elastography can provide elasticity information of PTMC quantitatively (VTQ) and directly reflects the overall elastic properties (VTI). Gender, hypoechogenicity, taller than wide, VTI elastography score ≥ 4, and SWV > 3.10 m/s are independent risk factors for predicting PTMC. ARFI elastography seems to be a new tool for the diagnosis of PTMC.

Solitary schwannoma of the gallbladder: a case report and literature review.

Schwannomas occurring in the gallbladder are extremely rare. Preoperative diagnosis of gallbladder schwannomas appears to be very difficult because they are normally asymptomatic and are often found incidentally. Until now, only five cases have been reported in the literature. To our knowledge, the contrast-enhanced ultrasound (CEUS) features of gallbladder schwannomas have not been reported before in other studies. We treated a 55-year-old male patient with gallbladder schwannoma in China. He had no symptoms, and the lesion was incidentally found by conventional ultrasound (US) when performing a health examination. The patient had normal liver function; moreover, serum carcinoembryonic antigen and alpha-fetoprotein were within the normal ranges. The lesion showed no blood flow signals on color Doppler US, and the wall beneath the lesion was intact on CEUS. The lesion was believed to be a benign entity; in addition, gallbladder adenomyomatosis was suspected. A laparoscopic cholecystectomy was performed to remove the mass. Pathological examination revealed that the tumor was mainly composed of spindle-shaped cells; neither atypical cells nor signs of malignancy were found. Immunohistochemical staining showed a strong positive S-100 protein reaction. Vimentin and CD56 staining were also positive, whereas CD34 and CD117 were negative. Finally, the lesion was diagnosed as schwannoma. Herein, we report the case; the associated literature is also reviewed.

The safety and treatment response of combination therapy of radioimmunotherapy and radiofrequency ablation for solid tumor: a study in vivo.

OBJECTION: To investigate the safety and treatment response of radioimmunotherapy (RIT) in combination with radiofrequency ablation (RFA) for the treatment of VX2 tumor on rabbit. MATERIALS AND METHODS: A total of 36 rabbits bearing VX2 tumor on the thigh were randomly assigned into 3 groups (group I: 1-2 cm; group II: 2-3 cm; group III: 3-4 cm) and 4 subgroups (A: as control, just puncture the tumor using the RFA electrode without power output; B: RFA alone; C: 131I-chTNT intratumoral injection alone; D: RFA+131I-chTNT intratumoral injection 3 days later). The variation of blood assay, weight and survival among different groups and subgroups were used to assess the treatment safety. Ultrasound (US) was used to monitor and assess the tumor response after treatment. RESULTS: According to the results of the weight and the blood assay among different groups, subgroups, and at two time points (one day before and the 16th day after treatment), no damages to the liver, kidney function and myelosuppression resulting from the treatment were found. No significant differences in survivals among the four subgroups (p = 0.087) were found. In addition, 131I-chTNT did not show significant inhibition effect on VX2 tumor progression according to US measurements. CONCLUSION: 131I-chTNT intratumoral injection alone or in combination with RFA is relatively safe for rabbit without significant toxicity and shows no significant effect on the survival. The treatment response is not as satisfactory as anticipated.

Conventional US, US elasticity imaging, and acoustic radiation force impulse imaging for prediction of malignancy in thyroid nodules.

PURPOSE: To evaluate conventional ultrasonography (US), US elasticity imaging (EI), and acoustic radiation force impulse (ARFI) imaging in thyroid nodule malignancy prediction. MATERIALS AND METHODS: This prospective study was institutional review board approved; informed consent was obtained. Study included 375 patients (mean age, 51 years; range, 18-75 years) with 441 pathologically proven thyroid nodules. In 281 women (mean age, 50 years; range, 18-75 years) and 94 men (mean age, 53 years; range, 18-74 years), conventional US, EI, Virtual Touch tissue imaging (VTi; Siemens, Mountain View, Calif), and Virtual Touch tissue quantification (VTq; Siemens) of ARFI imaging were performed for each nodule. Multivariate logistic regression analysis was performed to assess 17 independent variables for malignancy prediction. Diagnostic performance was evaluated with receiver operating characteristic (ROC) curve analysis. RESULTS: There were 325 benign and 116 malignant nodules. Marked hypoechogenicity (odds ratio [OR]: 83.88; 95% confidence interval [CI]: 17.81, 394.99) was the strongest independent predictor for thyroid malignancy, followed by shape taller than wide (OR: 8.69; 95% CI: 2.87, 26.31), VTi (OR: 6.54; 95% CI: 3.61, 11.88), moderate hypoechogenicity (OR: 3.98; 95% CI: 1.13, 14.05), poorly defined margin (OR: 3.27; 95% CI: 1.22, 8.77), female sex (OR: 2.55; 95% CI: 1.33, 4.91), coarse background of surrounding thyroid tissue (OR: 2.01; 95% CI: 1.12, 3.62), and VTq (OR: 1.78; 95% CI: 1.28, 2.47) (all P < .05). EI was not significantly associated with thyroid malignancy (P = .855). Area under the ROC curve (Az) for VTq and VTi was higher than that with other significant independent variables. Az, sensitivity, and specificity were 0.91 (95% CI: 0.87, 0.94) and 0.86 (95% CI: 0.82, 0.90), 80% and 71.6%, and 93.8% and 83.4%, respectively, for VTi and VTq. VTq of at least 2.87 m/sec and VTi of at least grade IV were the best cutoff values for malignant thyroid nodules. CONCLUSION: ARFI imaging is promising for malignant thyroid nodule prediction, with higher diagnostic performance than conventional US or EI. ARFI can be used to supplement conventional US to diagnose thyroid nodules in patients referred for surgery.

Sorafenib blocks the HIF-1α/VEGFA pathway, inhibits tumor invasion, and induces apoptosis in hepatoma cells.

Hypoxia and hypoxia-driven angiogenesis play an important role on the recurrence of hepatocellular carcinoma after insufficient radiofrequency ablation. The hypoxia-inducible factor (HIF)-1α/vascular endothelial growth factor-A (VEGFA) pathway plays an important part in this process. Sorafenib is a multikinase inhibitor with activity against several receptor tyrosine kinases. However, it is unclear whether sorafenib can affect the HIF-1α/VEGFA pathway. Here, we explore whether sorafenib affects HIF-1α and the change of invasion ability in this process. In this experiment, the control group, cobalt chloride (CoCl2)-treated group, sorafenib-treated group, and cobalt chloride combined with sorafenib-treated group were adopted. Western blot and PCR were performed to detect the protein and mRNA expression of HIF-1α and VEGFA in different groups. Transwell assay was used to test the changes of invasion ability. Flow cytometry was adopted to detect the apoptotic role of sorafenib on hepatoma cells. Cobalt chloride upregulated the expression of HIF-1α protein, and the upregulation effect was more obvious when the concentration was increased gradually. Sorafenib inhibited cobalt-induced HIF-1α and VEGFA expression in hepatoma cells. Sorafenib decreased the tumor cell invasiveness induced by cobalt chloride in vitro. Sorafenib inhibited cell proliferation and induced apoptosis in hepatoma cells. These results showed that sorafenib was an effective inhibitor of the HIF-1α/VEGFA pathway, which can provide new insight into the mechanism of its anticancer activity.

Management of hepatocellular carcinoma: The role of contrast-enhanced ultrasound.

Hepatocellular carcinoma (HCC) is the sixth most common neoplasm and the third cause of cancer death worldwide. Contrast enhanced ultrasound (CEUS) has been applied for more than ten years and plays increasingly important roles in the management of HCC. On the basis of the Guideline and Good Clinical Practice Recommendations for CEUS in the liver-update 2012 and related literature about the management of HCC, we summarize the main roles and applications of CEUS in the management of HCC, including HCC surveillance, diagnosis, CEUS-guided treatment, treatment response evaluation and follow-up. The diagnostic algorithm for HCC is also suggested. Meanwhile, the comparisons between CEUS and contrast enhanced computed tomography/magnetic resonance imaging (CECT/CEMRI) in these areas are made. Although CEUS is subject to the same limitation as ordinary US and is inferior to CECT/CEMRI in some aspects, CEUS has proved to be of great value in the management of HCC with inherent advantages, such as sufficient high safety profile making it suitable for patients with renal failure or allergic to iodine, absence of radiation, easy reproducibility and high temporal resolution. The tremendous application of CEUS to the diagnosis and treatment of HCC provides more opportunities for patients with HCC diagnosed at different stages.

Radiofrequency ablation before intratumoral injection of (131)I-chTNT improves the tumor-to-normal tissue ratio in solid VX2 tumor.

PURPOSE: This study was aimed to investigate whether the tumor necrosis induced by radiofrequency ablation (RFA) can improve the ratio of tumor-to-normal tissue (T/NT) after intratumoral injection of (131)I-chTNT. MATERIALS AND METHOD: Eighteen New Zealand rabbits bearing VX2 tumor on the thigh were randomly divided into two treatment groups (control group: intratumoral injection of (131)I-chTNT alone; RFA group: RFA + intratumoral injection of (131)I-chTNT 3 days after RFA) and each group was further divided into three subgroups I, II, and III (1-2 cm, 2-3 cm, and 3-4 cm in maximum diameter, respectively), by the tumor size. SPECT was performed to evaluate the T/NT on days 1, 8, and 15 after (131)I-chTNT injection. RESULTS: After treatment, all rabbits underwent the SPECT whole-body scan and the T/NT was analyzed. The results showed that T/NT in the RFA group (55.45±41.83) was significantly higher compared with the control group (7.23±5.61) (F=18.89, p=0.001). Meanwhile, a linear ascending trend was found for T/NT in the RFA group along with the follow-up time (r=0.47, p=0.01). The tumor size or the dose of (131)I-TNT injection had no significant effect on the variation of T/NT in both groups (p>0.05). CONCLUSION: RFA before intratumoral injection of (131)I-chTNT can dramatically improve T/NT, demonstrating the potential application of this combination therapy.

Role of contrast-enhanced ultrasound in follow-up assessment after ablation for hepatocellular carcinoma.

AIM: To assess the usefulness of contrast-enhanced ultrasound (CEUS) during follow-up after percutaneous ablation therapy for hepatocellular carcinoma (HCC). METHODS: A total of 141 patients with HCCs who received percutaneous ablation therapy were assessed by paired follow-up CEUS and contrast-enhanced computed tomography (CECT). The follow-up scheme was designed prospectively and the intervals between CEUS and CECT examinations were less than 14 d. Both images of follow-up CEUS and CECT were reviewed by radiologists. The ablated lesions were evaluated and classified as local tumor progression (LTP) and LTP-free. LTP was defined as regrowth of tumor inside or adjacent to the successfully treated nodule. The detected new intrahepatic recurrences were also evaluated and defined as presence of intrahepatic new foci. On CEUS and CECT, LTP and new intrahepatic recurrence both were displayed as typical enhancement pattern of HCC (i.e., hyper-enhancing during the arterial phase and washout in the late phase). With CECT as the reference standard, the ability of CEUS in detecting LTP or new intrahepatic recurrence during follow-up was evaluated. RESULTS: During a follow-up period of 1-31 mo (median, 4 mo), 169 paired CEUS and CECT examinations were carried out for the 141 patients. For a total of 221 ablated lesions, 266 comparisons between CEUS and CECT findings were performed. Thirty-three LTPs were detected on CEUS whereas 40 LTPs were detected on CECT, there was significant difference (P < 0.001). In comparison with CECT, the numbers of false positive and false negative LTPs detected on CEUS were 6 and 13, respectively; the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and overall accuracy of CEUS in detecting LTPs were 67.5%, 97.4%, 81.8%, 94.4% and 92.3%, respectively. Meanwhile, 131 new intrahepatic recurrent foci were detected on CEUS whereas 183 were detected on CECT, there was also significant difference (P < 0.05). In comparison with CECT, the numbers of false positive and false negative intrahepatic recurrences detected on CEUS were 13 and 65, respectively; the sensitivity, specificity, PPV, NPV and overall accuracy of CEUS in detecting new intrahepatic recurrent foci were 77.7%, 92.0%, 92.4%, 76.7% and 84.0%, respectively. CONCLUSION: The sensitivity of CEUS in detecting LTP and new intrahepatic recurrence after percutaneous ablation therapy is relatively low in comparison with CECT.

Contrast-enhanced ultrasound versus conventional ultrasound in the diagnosis of polypoid lesion of gallbladder: a multi-center study of dynamic microvascularization.

OBJECTIVE: To assess the value of contrast-enhanced ultrasound (CEUS) in the diagnosis of polypoid lesions of gallbladder (PLGs). METHODS: CEUS was performed to 116 patients (mean age, 49.6 years; range, 21-80 years) with PLGs from 8 university hospitals. 9 cases of biliary sludge were proven by surgery and the remaining 107 cases were confirmed by histopathological examination. The confidence level, diagnostic performance, inter-observer agreement of two independent readers with different experience was assessed. The readers were blind to the imaging and clinical results of the patients. RESULTS: There were significant differences between benign and malignant PLGs in patient age, gender, lesion size, echogenicity, stalk, time-to-peak, vascularity on CEUS, enhancement pattern, and wall destruction. The confidence levels increased significantly and the interobserver agreement increased from 0.425 to 0.601 after CEUS. The sensitivity increased from 22.2 to 77.8% after CEUS in the staff radiologist, and from 22.2 to 66.7% in the resident radiologist. The correctly characterized lesions were 64.7% before versus 87.1% after CEUS (P=0.125) for the staff radiologist, and 57.8% versus 70.7% for the resident radiologist (P=0.007). No significance was found in the subgroup of lesions≤1.0 cm before and after CEUS for the two radiologists. CONCLUSIONS: CEUS using convex multifrequency probes could detect the dynamic microvascularization of PLGs greater than 1.0 cm and facilitate the differentiation between benign and malignant tumors.

Parametric imaging with contrast-enhanced ultrasound: usefulness for characterization of dynamic effects of microvascularization for hepatocellular carcinoma and focal nodular hyperplasia.

OBJECTIVE: To evaluate whether parametric imaging with contrast-enhanced ultrasound (CEUS) is equal to experienced radiologists after review of CEUS in differentiating hepatocellular carcinoma (HCC) from focal nodular hyperplasia (FNH). METHODS: An image processing software was used to quantitatively analyze the CEUS clips of 30 HCCs (mean diameter, 3.4±0.9 cm; range, 1.8-5.0 cm) and 30 FNHs (mean diameter, 3.0±1.1 cm; range, 1.1-5.0 cm). Low mechanical index contrast specific imaging modes and contrast agent of SonoVue® were applied for CEUS. Fourteen HCCs were pathologically diagnosed and 16 were clinically diagnosed, whereas all the FNHs were confirmed by pathological examination. Quantitative parameters of HCC and FNH were compared. The diagnostic performance between parametric imaging and two experienced readers was compared using the receiver operating characteristic (ROC) curve analysis. RESULTS: On parametric imaging, the rise time, time to peak and mean transit time for HCC and FNH were 16.7±11.1 s vs. 21.9±9.0 s (P=0.052), 29.9±14.1 s vs. 33.2±11.1 s (P=0.322), 115.0±90.9 s vs. 271.5±147.6 s (P<0.001), respectively. The ROC analysis showed that, for the differentiation between HCC and FNH, the cut-off point for mTT was 107.93 s with the Az value of 0.817 (95% CI: 0.703-0.931), and the Az value was 0.834 (95%CI: 0.728-0.941) for two experienced readers (P=0.417 compared with mTT). The sensitivity, specificity, accuracy, positive predictive value (PPV), and negative predictive value (NPV) were 96.7%, 66.7%, 81.7%, 74.4%, and 95.2%, respectively, for parametric imaging, and 86.7%, 76.7%, 81.7%, 78.8%, and 85.2%, respectively, for two experienced readers (all P>0.05 compared with parametric imaging). CONCLUSION: Parametric imaging with CEUS is helpful for characterization the typical dynamic effects of microvascularization of HCC and FHH and is equal to experienced readers in the differential diagnosis between HCC and FNH.

Sorafenib suppresses the rapid progress of hepatocellular carcinoma after insufficient radiofrequency ablation therapy: an experiment in vivo.

BACKGROUND: Radiofrequency ablation (RFA) is a widely applied treatment for hepatocellular carcinoma (HCC), but insufficient RFA can promote rapid progression of the residual tumor through the hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor A (VEGFA) pathway. Although sorafenib has been successfully applied to advanced HCC, the use of sorafenib in residual tumor cells after RFA has rarely been tested. PURPOSE: To evaluate the potential role of sorafenib as an adjunct to RFA to reduce the recurrence rate after insufficient RFA. MATERIAL AND METHODS: Xenograft tumors of SMMC 7721 were created by subcutaneously inoculating nude mice with hepatoma cells (5 × 10(6) cells per mouse). Fourteen days after inoculation, all mice were divided into three groups (control group [sham puncture], RFA group, and RFA combined with sorafenib treatment group) with six mice in each group. Each group was given a different treatment procedure. After treatment, the volume of the tumors was calculated from the resected specimens. The mRNA and protein expression of HIF-1α and VEGFA was quantified by real-time PCR and immunohistochemistry analysis. The micro-vessel density (MVD) was determined by CD34 immunohistochemistry. RESULTS: Real-time PCR and immunohistochemistry analysis showed that, compared to the RFA group, HIF-1α and VEGFA expression were significantly decreased in the group that received RFA combined with sorafenib treatment (P < 0.05). By comparing the control group with the RFA group, we found that insufficient RFA promoted HIF-1α and VEGFA expression (P < 0.05). Similar results were obtained for MVD expression. Additionally, the combination of RFA with sorafenib therapy resulted in a synergistic reduction in tumor growth compared to insufficient RFA and sham puncture (P < 0.05). CONCLUSION: Sorafenib was able to inhibit the expression of HIF-1α and VEGFA, and sorafenib was able to increase time to recurrence when used as an adjunct to RFA.

Nano/microparticles and ultrasound contrast agents.

Microbubbles have been used for many years now in clinical practice as contrast agents in ultrasound imaging. Recently, their therapeutic applications have also attracted more attention. However, the short circulation time (minutes) and relatively large size (two to ten micrometers) of currently used commercial microbubbles do not allow effective extravasation into tumor tissue, preventing efficient tumor targeting. Fortunately, more multifunctional and theranostic nanoparticles with some special advantages over the traditional microbubbles have been widely investigated and explored for biomedical applications. The way to synthesize an ideal ultrasound contrast agent based on nanoparticles in order to achieve an expected effect on contrast imaging is a key technique. Currently a number of nanomaterials, including liposomes, polymers, micelles, dendrimers, emulsions, quantum dots, solid nanoparticles etc., have already been applied to pre or clinical trials. Multifunctional and theranostic nanoparticles with some special advantages, such as the tumor-targeted (passive or active), multi-mode contrast agents (magnetic resonance imaging, ultrasonography or fluorescence), carrier or enhancer of drug delivery, and combined chemo or thermal therapy etc., are rapidly gaining popularity and have shown a promising application in the field of cancer treatment. In this mini review, the trends and the advances of multifunctional and theranostic nanoparticles are briefly discussed.