Radiographic and α-fetoprotein response predict pathologic complete response to immunotherapy plus a TKI in hepatocellular carcinoma: a multicenter study.

BACKGROUND: Pathologic complete response (pCR) following preoperative systemic therapy is associated with improved outcomes after subsequent liver transplant/resection in hepatocellular carcinoma (HCC). However, the relationship between radiographic and histopathological response remains unclear. METHODS: We retrospectively examined patients with initially unresectable HCC who received tyrosine kinase inhibitor (TKI) plus anti-programmed death 1 (PD-1) therapy before undergoing liver resection between March 2019 and September 2021 across 7 hospitals in China. Radiographic response was evaluated using mRECIST. A pCR was defined as no viable tumor cells in resected samples. RESULTS: We included 35 eligible patients, of whom 15 (42.9%) achieved pCR after systemic therapy. After a median follow-up of 13.2 months, tumors recurred in 8 non-pCR and 1 pCR patient. Before resection, there were 6 complete responses, 24 partial responses, 4 stable disease cases, and 1 progressive disease case, per mRECIST. Predicting pCR by radiographic response yielded an area under the receiver operating characteristic curve (AUC) of 0.727 (95% CI: 0.558-0.902), with an optimal cutoff value of 80% reduction in the enhanced area in MRI (called major radiographic response), which had a 66.7% sensitivity, 85.0% specificity, and a 77.1% diagnostic accuracy. When radiographic response was combined with α-fetoprotein response, the AUC was 0.926 (95% CI: 0.785-0.999); the optimal cutoff value was 0.446, which had a 91.7% sensitivity, 84.6%, specificity, and an 88.0% diagnostic accuracy. CONCLUSIONS: In patients with unresectable HCC receiving combined TKI/anti-PD 1 therapy, major radiographic response alone or combined with α-fetoprotein response may predict pCR.

An alternative technique of arterial anastomosis in mouse heart transplantation.

OBJECTIVE: To investigate the characters and feasibility of continuous-interrupted suture (CIS) method for arterial anastomosis in mouse heart transplantation (MHT). METHODS: A MHT model was adopted. End-to-end anastomosis of donor ascending aorta to recipient abdominal aorta was achieved by CIS or continuous suture (CS) method. In both groups, end-to-end anastomosis of pulmonary vein to inferior vena cava (IVC) was achieved by CS. Technical indexes and histological examination were analyzed between 2 groups. RESULTS: The total operative time in CIS group was 92.83 ± 2.13 minutes, and in CS group was 92.40 ± 3.85 minutes. In CS group, artery anastomosis time was 20.13 ± 1.89 minutes; in CIS group, it was 20.36 ± 1.09 minutes. Additionally, venous anastomosis time in CS group was 14.80 ± 0.84 minutes, and in CIS group was 15.03 ± 0.85 minutes. Operation success rate in CIS group was 100%, and in CS group was 80%. There were no significant histological findings differences in graft between 2 groups. However, cell arrangement of anastomosis site was lightly irregular and the vascular alignment was poor in CS group. In CIS group, cell arrangement of anastomosis site was well arranged and vessels were well aligned. CONCLUSIONS: CIS method could avoid arterial anastomosis-related complications induced by CS and improve the success rate.

Circulating tumor cells are correlated with disease progression and treatment response in an orthotopic hepatocellular carcinoma model.

Hepatocellular carcinoma (HCC) is a highly malignant tumor characterized by rapid progression, poor prognosis, and frequent hematogenous metastasis. A minimally invasive diagnostic biomarker that can predict disease progression and treatment response would be of extraordinary benefit. Therefore, we have investigated whether the number of circulating tumor cells (CTCs) is correlated with disease progression and treatment response in HCC. Here we report that the number of CTCs, monitored by in vivo flow cytometry (IVFC), is strongly correlated with disease progression and treatment response in a highly metastatic orthotopic nude mouse model of green fluorescent protein (GFP)-labeled HCC. Sorafenib treatment reduces the number of CTCs significantly. The decreased number of CTCs is consistent with low lung metastasis. This study has demonstrated a considerable clinical value of CTCs as a biomarker in predicting disease progression and monitoring therapeutic efficacy in patients with HCC.

MiR-182 is up-regulated and targeting Cebpa in hepatocellular carcinoma.

MicroRNAs (miRNAs) are endogenous small non-coding RNAs that repress their targets at post transcriptional level. Existing studies have shown that miRNAs are important regulatory genes in hepatocellular carcinoma (HCC), as either tumor suppressors or oncogenes. MiR-122 is normally downregulated in HCC and regarded as a tumor suppressor. Recently miR-122 has been reported to be regulated by CEBPA, which is then involved in a novel pathway to influence proliferation of tumor cells. However it is unknown whether CEBPA is regulated by miRNAs in HCC. In this study, we find that miR-182 is upregulated in HCC model rat, and represses CEBPA in both rat and human. This further improves the current CEBPA/miR-122 pathway that controls the proliferation of tumor cells. These results suggest that miR-182 is a potential oncogene in HCC and could be used as a diagnostic marker and drug target of HCC.

Sorafenib delays recurrence and metastasis after liver transplantation in a rat model of hepatocellular carcinoma with high expression of phosphorylated extracellular signal-regulated kinase.

Liver transplantation (LT) is one of the curative treatments for hepatocellular carcinoma (HCC). However, cancer recurrence and metastasis after LT are common in some HCC patients with high-risk factors (even in those within the Milan criteria). It remains unclear whether adjuvant therapy with sorafenib inhibits HCC recurrence and metastasis after LT. Therefore, we performed orthotopic LT in an August Irish Copenhagen (ACI) rat model of HCC. Because LT involves immune rejection and tolerance and it is unknown whether sorafenib influences the immune response, we also investigated the effects of sorafenib on immune balance. In this study, we established an allogeneic rat LT model in which liver grafts were taken from Lewis rats and transplanted into ACI rats with orthotopic HCC, and they were administered cyclosporine A to prevent acute allograft rejection. From day 7 after LT, sorafenib was administrated at 30 mg/kg/day for 3 weeks. Our results showed that the serum levels of vascular endothelial growth factor and hepatocyte growth factor significantly increased after LT, and the T helper 1 (T(h)1)/T helper 2 (T(h)2) immune balance was shifted toward a T(h)2 response after immunosuppressant administration. In comparison with controls, the rats in the sorafenib group showed significantly inhibited extracellular signal-regulated kinase phosphorylation and improved progression-free survival and overall survival. The tumor proliferation rate and angiogenesis in posttransplant recurrent tumor tissues decreased in the sorafenib group, and the tumor apoptosis rate increased. There was no significant difference in the T(h)1/T(h)2 immune balance between the sorafenib and control groups. In conclusion, adjuvant therapy with sorafenib is highly effective at inhibiting cancer recurrence and metastasis without influencing the immune balance after LT for HCC with high expression of phosphorylated extracellular signal-regulated kinase. This study suggests that sorafenib may have potential, particularly as part of a stratified medicine approach to HCC treatment after LT.

Effectiveness and safety of lenvatinib plus anti-programmed death-1 antibodies in patients with hepatocellular carcinoma: A real-world cohort study.

OBJECTIVE: Lenvatinib plus anti-programmed death-1 (anti-PD-1) antibody combinations have shown potent anti-tumor effect in phase I/II trials in advanced or unresectable hepatocellular carcinoma (HCC), but real-world data are limited. METHODS: To investigate the effectiveness and safety of lenvatinib plus anti-PD-1 antibodies in a real-world cohort, we retrospectively evaluated 210 patients with unresectable or advanced HCC treated with these regimens between October 2018 and February 2022. RESULTS: The objective response rate and disease control rate per Response Evaluation Criteria in Solid Tumors (RECIST) v1.1 were 28.1% and 75.2%. Median overall survival (OS) and progression-free survival (PFS) in the overall cohort were 17.2 and 8.4 months, respectively. Median OS and PFS of patients receiving first-line treatment reached 18.9 and 9.6 months. Median OS was significantly longer in patients with Child-Pugh class A versus B (18.8 vs. 5.9 months, respectively), as was median PFS (9.1 vs. 4.4 months). Patients with albumin-bilirubin (ALBI) grade 1 versus grade 2/3 also had significantly greater median OS (23.5 vs. 13.4 months). Treatment-related adverse events (AEs) occurred in 79.5% of patients. Patients with ALBI grade 2/3 had a higher rate of grade 3/4 AEs than patients with ALBI grade 1 (57.5% vs. 38.5%). CONCLUSION: Lenvatinib combined with anti-PD-1 antibody therapy was effective in patients with sufficient liver function reserve. Further study is needed to improve therapeutic efficacy and AE management in patients with Child-Pugh class B or ALBI grade 2/3.

The Safety Profile of Hepatectomy Following Preoperative Systemic Therapy with Lenvatinib Plus Anti-PD-1 Antibodies Versus Hepatectomy Alone in Patients With Hepatocellular Carcinoma.

Objective: To determine the safety of hepatectomy after combined lenvatinib and anti-PD-1 preoperative systemic therapy (PST) in patients with marginally resectable hepatocellular carcinoma (HCC). Background: PST followed by hepatectomy (PSTH) is an emerging treatment for HCC. However, the impact of PST with lenvatinib plus anti-PD-1 antibodies on surgical safety is unknown. Methods: Medical records from consecutive patients with marginally resectable advanced HCC who underwent hepatectomy after PST with lenvatinib and anti-PD-1 antibodies between January 2018 and August 2021 were retrieved from a prospectively designed database. Propensity score matching (1:2) was performed with a further 2318 HCC patients who underwent upfront hepatectomy (UH) without initial antitumor treatment during the same period. Results: In total, 49 and 98 matched patients were included in the PSTH and UH groups, respectively. Compared to the UH group, individuals in the PSTH group experienced more intraoperative blood loss, blood transfusions, and longer postoperative hospital stays. Moreover, posthepatectomy liver failure was more common in the PSTH group, who also had worse albumin-bilirubin (ALBI) scores on postoperative days 1-7. A significantly greater amount of drainage was also required in the PSTH group. However, the 30-day morbidity and 90-day mortality were similar among the two groups. Additionally, the duration of surgery, use of hepatic inflow occlusion during surgery, and the levels of postoperative inflammation-based markers were not statistically different between the two groups. Conclusions: Despite more intraoperative and postoperative adverse events, PSTH had comparable 30-day morbidity and 90-day mortality as UH. Thus, PSTH appears to be a viable treatment option for marginally resectable HCC patients with careful preoperative evaluation.

Organ specific responses to first-line lenvatinib plus anti-PD-1 antibodies in patients with unresectable hepatocellular carcinoma: a retrospective analysis.

BACKGROUND: We evaluated organ-specific response rates (OSRRs) to first-line lenvatinib plus anti-PD-1 antibodies in patients with advanced hepatocellular carcinoma (HCC). METHODS: This retrospective analysis included Chinese patients with unresectable/advanced HCC who received first-line lenvatinib (8 mg/day) plus ≥3 infusions of anti-PD-1 antibodies between October 2018 and May 2020. Tumor and macrovascular tumor thrombi (MVTT) treatment responses were evaluated every 2 months using RECIST v1.1. The overall response rate (ORR)/OSRR was defined as the percentage of patients with a best overall response of complete or partial response (CR or PR). RESULTS: In total, 60 patients were included in the analysis; 96.7% had measurable intrahepatic lesions, 55% had MVTT and 26.7% had extrahepatic disease. In all 60 patients, the ORR was 33.3%, median progression-free survival was 7.0 months (95% CI, 1.7-12.3) and median overall survival was not reached. The OSRR for MVTT (54.5%) was higher versus intrahepatic tumors (32.8%), extrahepatic lung metastases (37.5%) and lymph node metastases (33.3%). Among 33 patients with intrahepatic tumors and MVTT, 18 had differential responses in each site, including 13 with a better response in MVTT versus intrahepatic lesions. Among 18 patients whose MVTT achieved a radiographic CR or PR, six underwent surgical resection: 4/6 achieved a pathological CR in MVTT and 2/6 in the intrahepatic tumor. CONCLUSIONS: First-line lenvatinib plus anti-PD-1 antibodies resulted in better tumor responses in MVTT versus intrahepatic lesions. Complete MVTT necrosis may allow downstaging and subsequent eligibility for surgical resection in a proportion of patients with advanced HCC.

Downstaging and Resection of Initially Unresectable Hepatocellular Carcinoma with Tyrosine Kinase Inhibitor and Anti-PD-1 Antibody Combinations.

BACKGROUND: Combined therapy with tyrosine kinase inhibitors (TKIs) and anti-PD-1 antibodies has shown high tumor response rates for patients with unresectable hepatocellular carcinoma (HCC). However, using this treatment strategy to convert initially unresectable HCC to resectable HCC was not reported. METHODS: Consecutive patients with unresectable HCC who received first-line therapy with combined TKI/anti-PD-1 antibodies were analyzed. Tumor response and resectability were evaluated via imaging every 2 months (±2 weeks) using RECIST v1.1. Resectability criteria were (1) R0 resection could be achieved with sufficient remnant liver volume and function; (2) intrahepatic lesions were evaluated as partial responses or stable disease for at least 2 months; (3) no severe or persistent adverse effects occurred; and (4) hepatectomy was not contraindicated. RESULTS: Sixty-three consecutive patients were enrolled. Of them, 10 (15.9%) underwent R0 resection in 3.2 months (range: 2.4-8.3 months) after the initiation of combination therapy. At baseline, these 10 patients had a median largest tumor diameter of 9.3 cm, 7 had Barcelona Clinic Liver Cancer stage C (vascular invasion) disease, 2 had stage B, and 1 had stage A. Before surgery, 6 patients were evaluated as a partial response, 3 stable disease, and 1 partial response in the intrahepatic lesion but a new metastatic lesion in the right adrenal gland. Six patients (60%) achieved a pathological complete response. One patient died from immune-related adverse effects 2.4 months after hepatectomy. After a median follow-up of 11.2 months (range: 7.8-15.9 months) for other 9 patients, 8 survived without disease recurrence, and 1 experienced tumor recurrence. CONCLUSIONS: Combination of TKI/anti-PD-1 antibodies is a feasible conversion therapy for patients with unresectable HCC to become resectable. This study represents the largest patient cohort on downstaging role of combinational systemic therapy on TKI and PD-1 antibody for HCC.

Capn4 overexpression underlies tumor invasion and metastasis after liver transplantation for hepatocellular carcinoma.

UNLABELLED: Liver transplantation (LT) is one of the best therapeutic options for nonresectable hepatocellular carcinoma (HCC). Unfortunately, some HCC patients succumb to the disease after LT, which reduces long- and medium-term survival. To identify the proteins associated with HCC invasion and metastasis, HCC patients undergoing LT with complete follow-up data were included in this study and were categorized into recurrence and nonrecurrence groups. We extracted the total protein from the acquired homogeneous tumor cells and applied a cleavable isotope-coded affinity tag technology to quantitate relative changes in protein levels between the two groups. We identified a total of 149 proteins with two-dimensional liquid chromatography coupled with tandem mass spectrometry, including 52 differentially expressed proteins by at least two-fold. Among them, calpain small subunit 1 (Capn4), a protein with relevant interactions with many migration-invasion-related proteins, has attracted more attention. First, Capn4 overexpression in the recurrence group was confirmed via real-time polymerase chain reaction and western blotting in another cohort of 40 HCC patients undergoing LT. Second, Capn4 was associated with enhanced invasiveness in vitro. The small interfering RNA-mediated knockdown expression of Capn4 in HCC cell lines significantly inhibited its mobile and invasive ability. Tissue microarray in a further 192 cases revealed that Capn4 significantly correlated with invasive phenotype of HCC, and univariate and multivariate analyses indicated that Capn4 is an independent prognostic factor for recurrence and survival of HCC patients. CONCLUSION: Our study revealed that Capn4 overexpression underlies invasion and metastasis after LT for HCC and might be a candidate biomarker for future diagnosis and a target for therapy.

Preventive effects of curcumin and dexamethasone on lung transplantation-associated lung injury in rats.

OBJECTIVE: To investigate potential effects of curcumin or dexamethasone on lung transplantation-associated lung injury. DESIGN: Prospective, randomized, controlled study. SETTING: Research laboratory. SUBJECTS: Adult male Sprague-Dawley rats. INTERVENTIONS: Sham-operated rats were used as time-matched controls. Experimental rats were subjected to unilateral orthotopic lung transplantation with 4 hrs of cold ischemia followed by 2 hrs (or 24 hrs) of reperfusion. Animals were randomly assigned to vehicle-, curcumin-, or dexamethasone-treated groups. MEASUREMENTS AND MAIN RESULTS: Transplantation-associated lung injury was characterized by an increased alveolar-capillary permeability and myeloperoxidase activity and decreased levels of arterial oxygen tension/inspired oxygen concentration ratio. Pretreatment with curcumin and dexamethasone significantly prevented barrier disruption, lung edema, tissue inflammation, and decreased PaO2 at the early stage of posttransplantation. Nuclear factor-kappaB in transplanted lungs was activated, accompanied by an increase in messenger RNA levels and protein content of tumor necrosis factor-alpha, interleukin-6, and matrix metalloproteinase-9 in lung graft. Those changes were prevented by pretreatment with curcumin and dexamethasone. CONCLUSIONS: Curcumin can be an alternative therapy for protecting lung transplantation-associated injury by suppressing nuclear factor-kappaB-mediated expression of inflammatory genes.

Pedigree-based genome re-sequencing reveals genetic variation patterns of elite backbone varieties during modern rice improvement.

Rice breeding has achieved great productivity improvements by semi-dwarf varieties and hybrid vigour. Due to poor understanding of genetic basis of elite backbone varieties, the continuous increasing in rice yield still faces great challenges. Here, 52 elite rice varieties from three historical representative pedigrees were re-sequenced with 10.1× depth on average, and ~6.5 million single nucleotide polymorphisms (SNPs) were obtained. We identified thousands of low-diversity genomic regions and 0-diversity genes during breeding. Using pedigree information, we also traced SNP transmission patterns and observed breeding signatures in pedigree. These regions included the larger number of key well-known functional genes. Besides, 35 regions spanning 0.16% of the rice gnome had been differentially selected between conventional and restorer pedigrees. These genes identified here will be useful to the further pedigree breeding. Our study provides insights into the genetic basis of backbone varieties and will have immediate implications for performing genome-wide breeding by design.

Factors influencing survival in hepatocellular carcinoma patients with macroscopic portal vein tumor thrombosis after surgery, with special reference to time dependency: a single-center experience of 381 cases.

BACKGROUND/AIMS: The prognosis ofhepatocellular carcinoma with macroscopic portal vein tumor thrombosis is extremely poor. The risk factors may differ at different postoperative intervals. This study was undertaken to clarify the surgical outcome and time dependency of factors influencing survival in these patients. METHODOLOGY: We analyzed clinicopathological variables of 381 hepatocellular carcinoma patients with macroscopic portal vein tumor thrombosis who underwent hepatic resection. Survival rates were calculated using Kaplan-Meier method. The stratified Cox models were used to identify factors independently influencing short- and long-term survival, respectively. RESULTS: The cumulative 1-, 2-, 3-, 5-, and 10-year survival rates in 381 patients were 47%, 23%, 16%, 12%, 6%, respectively. The 1-, 3-, and 5-year survival rates calculated from time of re-resection were 36%, 14% and 0% in patients undergoing re-resection for intrahepatic recurrence within 2 years after first operation, and 85%, 53% and 32% in those more than 2 years after first operation (P<0.05). Multivariate analysis showed that portal vein infusion chemotherapy, serum alpha-fetoprotein > 20 mg/L and positive surgical margin were significant prognostic factors within 2 years after operation. In contrast, alanine aminotransferase > 80 U/L was the only significant factor beyond 2 years after operation. CONCLUSIONS: The survival of hepatocellular carcinoma patients with macroscopic portal vein tumor thrombosis was poor, but the prognosis of patients who had tumor recurrence more than 2 years after operation was much better than those with tumor recurrence within 2 years. Evaluation of time-dependency of risk factors may have important clinical implication in determining the therapeutic strategy.

[Time dependency of factors influencing survival of hepatocellular carcinoma patients with portal vein tumor thrombosis after surgery].

OBJECTIVE: To investigate the surgical outcome of the hepatocellular carcinoma (HCC) patients with portal vein tumor thrombosis (PVTT) after surgery and the time-dependency of the factors influencing survival. METHODS: The clinicopathological data of 382 HCC patients with macroscopic PVTT who had undergone resection of HCC were analyzed. The survival rte was calculated using Kaplan-Meier method. Stratified Cox model was used to identify the factors independently influencing the short- and long-term survival rates. RESULTS: The 1-, 2-, 3-, 5-, and 10-year survival rates of the 382 patients were 47%, 23%, 16%, 12%, and 6% respectively. The 1-, 3-, and 5-year survival rates re-calculated from the time of re-resection because of recurrence within 2 years after the first operation were 36%, 14%, and 0% 1 respectively. However, the 1-, 3-, and 5-year survival rates re-calculated from the time of re-resection because of recurrence 2 years after the first operation were 85%, 53%, and 32%, all significantly higher than those re-calculated from the time of re-resection within 2 years after the first operation (all P < 0.05). Multivariate analysis showed that portal infusion chemotherapy, serum alpha-fetoprotein < 20 microg/L and negative surgical margin were significant favorable prognostic factors within 2 years after operation. Alanine aminotransferase > 80 U/L was the only significant unfavorable factor beyond 2 years after operation. CONCLUSION: The prognosis of the patients with macroscopic PVTT who suffer from liver tumor recurrence occurring more than 2 years after the first operation is much better than those with the recurrence occurring within 2 years. Evaluation of the time-dependency of risk factors may have important clinical implication in determining the therapeutic strategy.

Tacrolimus promotes hepatocellular carcinoma and enhances CXCR4/SDF­1α expression in vivo.

The aim of our study was to elucidate the effect of tacrolimus (FK506) and of C-X-C chemokine receptor type 4 (CXCR4), which is a receptor specific to the stromal cell-derived factor-1α (SDF­1α), on growth and metastasis of hepatocellular carcinoma (HCC). Following treatment with different concentrations of FK506, AMD3100 or normal saline (NS), the proliferation of Morris rat hepatoma 3924A (MH3924A) cells was measured by the MTT assay, the expression of CXCR4 was analyzed with immunohistochemistry, and the morphological changes and the invasiveness of cells were studied with a transwell assay and under a scanning electron microscope, respectively. In addition, August Copenhagen Irish rat models implanted with tumor were used to examine the pathological changes and invasiveness of tumor in vivo, the expression of CXCR4 in tumor tissues and the expression of SDF­1α in the adjacent tissues to the HCC ones, using immunohistochemistry. In vitro, FK506 (100­1,000 µg/l) significantly promoted the proliferation of MH3924A cells (P<0.01), and increased the expression of CXCR4 in MH3924A cells, albeit with no significance (P>0.05). By contrast, AMD3100 had no effect on the proliferation of MH3924A cells, but significantly reduced the expression of CXCR4 (P<0.05). The invasiveness of MH3924A cells was significantly (P<0.01) enhanced following treatment with FK506, SDF­1α, FK506 + AMD3100, FK506 + SDF­1α or FK506 + AMD3100 + SDF­1α. In vivo, tumor weight (P=0.041), lymph node metastasis (P=0.002), the number of pulmonary nodules (P=0.012), the expression of CXCR4 in tumor tissues (P=0.048) and that of SDF­1α in adjacent tissues (P=0.026) were significantly different between the FK506-treated and the NS group. Our results suggest that FK506 promotes the proliferation of MH3924A cells and the expression of CXCR4 and SDF­1α in vivo. Therefore, inhibiting the formation of the CXCR4/SDF­1α complex may partly reduce the promoting effect of FK506 on HCC.

Expression of hemopexin in acute rejection of rat liver allograft identified by serum proteomic analysis.

Acute rejection (AR) and acceptance of allograft after liver transplantation (LTx) remain critical issues that need addressing to improve prognosis. We therefore performed rat orthotopic LTx and proteomic analyses to screen for immune response-related biomarkers in sera. Markers identified were validated at the mRNA and/or protein levels, and the molecules of interest were functionally explored. Compared with syngeneic controls, signs of AR as well as spontaneous acceptance were observed in hematoxylin and eosin-stained sections of liver allografts. In accordance with the severity of AR, 30 protein spots displaying significant changes in abundance were identified using two-dimensional differential gel electrophoresis. Ultimately, 14 serum proteins were sequenced and five spots of interest were identified as hemopexin (HPX). Expression of HPX was significantly and inversely associated with the severity of AR at both the mRNA and protein levels. In vitro, Mt-1, Ho-1, Fth, Ifn-γ, and Il-17 transcripts were significantly upregulated in lysates of lymphocytes stimulated with HPX, whereas Il-10 markedly was remarkably downregulated. Interferon-γ, IL-10, and IL-17 proteins in the supernatant of HPX-stimulated lymphocytes were significantly altered in keeping with the mRNA level. Our data facilitated the generation of a proteomic profile to enhance the understanding of rat liver AR. In view of finding that the HPX serum level is negatively associated with the severity of AR of rat liver allograft, we propose that in vitro treatment with HPX regulates cytokine expression in rat lymphocytes.

Plasma microRNA, a potential biomarker for acute rejection after liver transplantation.

BACKGROUND: Acute rejection (AR) of an organ transplant is a life-threatening complication. Currently, there are few diagnostic biomarkers suitable for clinical application. We aim to determine the potential of plasma microRNAs as biomarkers for AR. METHODS: Using rat orthotopic liver transplantation model and microarrays, we compared the difference in the spectrum and levels of microRNAs in both plasma and grafts between AR rats and control. AR-related plasma microRNAs were selected and validated using real-time quantification polymerase chain reaction. Plasma from AR rats with or without tacrolimus treatment was used for microRNA dynamic monitoring. To clarify the origin of AR-related plasma microRNAs, drug-induced liver damage rat model were performed and in situ hybridization was used to detect and localize the specific microRNA in allografts. RESULTS: We found that plasma miR-122, miR-192, and miR-146a was significantly up-regulated when AR occur (fold change>2; P<0.05) and the elevation could be repressed by immunosuppression. In liver injury rat model, up-regulated plasma miR-122 (fold change=22.126; P=0.002) and miR-192 (fold change=8.833; P<0.001) rather than miR-146a (fold change=1.181; P=0.594) were observed. Further study demonstrated that miR-146a was up-regulated by sixfold in microvesicles isolated from AR plasma, whereas miR-122 and miR-192 showed no distinct change. In situ hybridization revealed that the portal areas of the AR graft were brimming with lymphocytes, which showed highly intense staining for miR-146a. CONCLUSIONS: Our study provides the global fingerprint of plasma microRNAs in AR rats and suggests that plasma miR-122 and miR-192 reflect liver injury, whereas miR-146a may associate with cellular rejection.

Use of multiphoton microscopy to diagnose liver cancer and lung metastasis in an orthotopic rat model.

Liver or lung biopsy for suspicious lesions has several disadvantages such as bleeding, bile leak or pneumothorax, needle track seeding, and time-consuming histopathological procedure. The ability to directly observe cellular and subcellular details and then perform "optical biopsy" is a major goal in the development of new interventional techniques. Multiphoton microscopy (MPM) enables real-time noninvasive visualization of tissue architecture and cell morphology in live tissue. We performed a study to evaluate whether MPMcan make real-time optical diagnosis for liver cancer and lung metastasis using an orthotopic rat model with Morris hepatoma. We found that real-time high-resolution MPMimaging could clearly show tissue architecture and cell morphology. In the normal liver tissue, MPMimaging clearly revealed the blood-filled sinusoids and cords of hepatocytes. In the cancerous tissue, MPMimaging clearly illustrated that cancer cells displayed marked cellular and nuclear pleomorphism. MPMimages were comparable to golden standard hematoxylin-eosin staining images. Moreover, MPMimaging had deep penetration with the capability of optical sectioning. In short, MPMcan make real-time optical diagnosis for liver cancer and lung metastasis. This study provides the groundwork for further using multiphoton endoscopy to perform real-time noninvasive "optical biopsy" for liver cancer and lung metastasis in the near future.

Activation of PI3K/AKT and MAPK pathway through a PDGFRß-dependent feedback loop is involved in rapamycin resistance in hepatocellular carcinoma.

BACKGROUND: Rapamycin is an attractive approach for the treatment and prevention of HCC recurrence after liver transplantation. However, the objective response rates of rapamycin achieved with single-agent therapy were modest, supporting that rapamycin resistance is a frequently observed characteristic of many cancers. Some studies have been devoted to understanding the mechanisms of rapamycin resistance, however, the mechanisms are cell-type-dependent and studies on rapamycin resistance in HCC are extremely limited. METHODOLOGY/PRINCIPAL FINDINGS: The anti-tumor sensitivity of rapamycin was modest in vitro and in vivo. In both human and rat HCC cells, rapamycin up-regulated the expression and phosphorylation of PDGFRß in a time and dose-dependent manner as assessed by RT-PCR and western blot analysis. Using siRNA mediated knockdown of PDGFRß, we confirmed that subsequent activation of AKT and ERK was PDGFRß-dependent and compromised the anti-tumor activity of rapamycin. Then, blockade of this PDGFRß-dependent feedback loop by sorafenib enhanced the anti-tumor sensitivity of rapamycin in vitro and in an immunocompetent orthotopic rat model of HCC. CONCLUSIONS: Activation of PI3K/AKT and MAPK pathway through a PDGFRß-dependent feedback loop compromises the anti-tumor activity of rapamycin in HCC, and blockade of this feedback loop by sorafenib is an attractive approach to improve the anti-tumor effect of rapamycin, particularly in preventing or treating HCC recurrence after liver transplantation.

An investigation of the effect of sorafenib on tumour growth and recurrence after liver cancer resection in nude mice independent of phosphorylated extracellular signal-regulated kinase levels.

OBJECTIVE: The goal of this study is to investigate the effects of sorafenib on tumor growth, recurrence and metastasis after curative resection of liver cancer. RESEARCH METHODS: SMMC-7721 and HCCLM3 liver tumors, each with different metastatic potential and basal phosphorylated extracellular signal-regulated kinase (pERK) levels, were orthotopically implanted into 56 nude mice. Mice were divided into a treatment sub study and a prevention sub study. RESULTS: In the treatment sub study, tumor volumes in the high pERK-expressing HCCLM3 model were 2.58 ? 0.83 and 0.38 ? 0.09 cm(3) without and with sorafenib, respectively (p < 0.001). The corresponding volumes in the low pERK-expressing SMMC-7721 model were 1.36 ? 0.24 and 0.24 ? 0.14 cm(3) (p < 0.001), respectively. Sorafenib inhibited HCCLM3 cell proliferation and decreased tumor angiogenesis, but did not inhibit proliferation in the SMMC-7721 model. In the prevention sub study, intrahepatic recurrent tumor volumes were 1.96 ? 0.45 and 0.18 ? 0.24 cm(3) (p < 0.001); lung metastasis frequencies were 100 and 0% (p = 0.005); and lifespans were 36 ? 3 and 46 ? 5 days (p = 0.002) in the control and sorafenib subgroups, respectively. CONCLUSIONS: Sorafenib inhibits tumor growth and prevents metastatic recurrence after resection of hepatocellular carcinoma in nude mice. The effect of sorafenib does not exclusively depend on high levels of pERK in tumors.