Utility of sonic hedgehog and keratin 8/18 immunohistochemistry for detecting ballooned hepatocytes.

AIMS: Ballooned hepatocytes represent liver cell degeneration and are histological hallmarks in the diagnosis of non-alcoholic steatohepatitis, a severe form of non-alcoholic fatty liver disease. However, the identification of ballooned hepatocytes is often difficult, especially in the clinical setting of patients with other chronic liver diseases. In this study, we investigated the utility of immunostaining for positive sonic hedgehog (SHh) protein and negative Keratin 8/18 (K8/18) expression on ballooned hepatocytes. METHODS AND RESULTS: Immunohistochemistry for SHh and K8/18 was evaluated independently by two experienced liver pathologists in non-tumorous liver tissue from 100 cases of resected hepatocellular carcinoma of various aetiology. The degree of hepatocyte ballooning was scored as follows: 0, none; 1, few; 2, many ballooned hepatocytes. These evaluations were performed using routine haematoxylin and eosin (H&E) staining, followed by immunostaining for SHh or K8/18. Using SHh or K8/18 immunostaining combined with H&E staining, the score of ballooned hepatocytes was upgraded in 20 and 19 cases, and downgraded in none and 2 cases, respectively. The percentage of observed agreement for ballooned hepatocytes scoring was 85% and 92%, and the weighted kappa value was 0.806 and 0.893 with SHh or K8/18 immunohistochemistry. Considering the immunohistochemistry results, background liver disease diagnosis was changed in 15 out of 100 cases (15%) evaluated. CONCLUSIONS: SHh and K8/18 immunohistochemistry are useful in detecting ballooned hepatocytes, regardless of background liver disease, and improving pathological diagnosis accuracy.

Terminal cationization of poly(N-isopropylacrylamide) brush surfaces facilitates efficient thermoresponsive control of cell adhesion and detachment.

A variety of poly(N-isopropylacrylamide) (PIPAAm)-grafted surfaces have been reported for temperature-controlled cell adhesion/detachment. However, the surfaces reported to date need further improvement to achieve good outcomes for both cell adhesion and detachment, which are inherently contradictory behaviors. This study investigated the effects of terminal cationization and length of grafted PIPAAm chains on temperature-dependent cell behavior. PIPAAm brushes with three chain lengths were constructed on glass coverslips via surface-initiated reversible addition-fragmentation chain transfer (RAFT) polymerization. Terminal substitution of the grafted PIPAAm chains with either monocationic trimethylammonium or nonionic isopropyl moieties was performed through the reduction of terminal RAFT-related groups and subsequent thiol-ene reaction with the corresponding acrylamide derivatives. Although the thermoresponsive properties of the PIPAAm brush surfaces were scarcely affected by the terminal functional moiety, the zeta potentials of the cationized PIPAAm surfaces were higher than those of the nonionized ones, both below and above the phase transition temperature of PIPAAm (30°C). When bovine endothelial cells were cultured on each surface at 37°C, the number of adherent cells decreased with longer PIPAAm. Notably, cell adhesion on the cationized PIPAAm surfaces was higher than that on the nonionized surfaces. This terminal effect on cell adhesion gradually weakened with increasing PIPAAm length. In particular, long-chain PIPAAm brushes virtually showed cell repellency even at 37°C, regardless of the termini. Interestingly, moderately long-chain PIPAAm brushes promoted cell detachment at 20°C, with negligible terminal electrostatic interruption. Consequently, both cell adhesion and detachment were successfully improved by choosing an appropriate PIPAAm length with terminal cationization.

Pathological findings that contribute to tissue stiffness in the spleen of liver cirrhosis patients.

AIM: Spleen stiffness is increased in liver cirrhosis (LC). We attempted to characterize the pathological features of spleen in LC. METHODS: We compared pathological findings of resected spleen tissues of 28 LC patients and those of six healthy controls. In addition, we measured spleen stiffness before splenectomy by shear wave elastography in nine LC patients. After splenectomy, we examined the relationship between spleen stiffness and pathological findings. RESULTS: Passive congestion of the spleen was more frequently observed in LC patients than in controls (P < 0.01). The sinus was wider in LC patients than in controls (P < 0.01). In the spleens of the LC patients, diffuse α-smooth muscle actin (αSMA) expression of sinusoidal mesenchymal cells and deposition of collagen fibers on the perisinusoidal wall were observed. In nine LC patients whose spleen stiffness was examined, the width of the sinus increased along with spleen stiffness (r = 0.89, P < 0.01). Spleen stiffness was higher in the spleen tissues with diffuse αSMA expression of sinusoidal mesenchymal cells than in those with partial αSMA expression of sinusoidal mesenchymal cells (P = 0.01). The degree of fibrosis was higher in the LC patients with diffuse αSMA expression of the red pulp than in those with partial αSMA expression of the red pulp (P = 0.03). CONCLUSION: In the LC patients, spleen tissues showed passive congestion with a dilated sinus, diffuse αSMA expression of sinusoidal mesenchymal cells, and deposition of collagen fibers on the perisinusoidal wall. This contributed to spleen stiffness.

SUOX is negatively associated with multistep carcinogenesis and proliferation in oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is the most common malignant tumor in the head and neck region. The aim of this study was to identify the key molecules and to elucidate the molecular mechanisms of OSCC carcinogenesis through a microarray analysis of RNA extracted from normal epithelium, dysplasia, and squamous cell carcinoma components. Out of molecules that showed changes in gene expression in the microarray analysis, we focused on Sulfite oxidase (SUOX), which correlated significantly with carcinogenic process and exhibited a stepwise decrease in expression. The expression of SUOX was evaluated in detail at the protein level using samples from 58 patients with cancer of the tongue, and correlating clinicopathological factors were also comprehensively examined. SUOX expression declined significantly from normal epithelium to dysplasia to squamous cell carcinoma components in line with carcinogenic process. With regard to squamous cell carcinoma, SUOX expression was significantly lower when T classification was high. Our findings indicated that SUOX is negatively associated with the progression and proliferation of tongue cancer, and suggest that SUOX may be a key molecule in tongue tumors.

Insulinoma-associated protein 1 (INSM1) is a useful marker for pancreatic neuroendocrine tumor.

Insulinoma-associated protein 1 (INSM1) is an important biomarker of Achaete-scute homolog-like 1-driven pathways. For diagnosis of pancreatic neuroendocrine tumors (PanNET), chromogranin A (CGA), synaptophysin (SYP), and neural cell adhesion molecule (NCAM) were also considered as potential biomarkers. However, it is often difficult to diagnose it immunohistochemically. Hence, we examined the expression pattern of INSM1 in pancreatic solid tumors. We detected INSM1, CGA, SYP, and NCAM immunohistochemically, in 27 cases of NET [pure type: 25 cases, mixed adenoneuroendocrine carcinoma (MANEC): 2 cases]. We included 5 cases of solid-pseudopapillary neoplasm (SPN), 7 cases of acinar cell carcinoma (ACC), and 15 cases of pancreatic ductal adenocarcinoma (PDAC) as the control group. Nuclear expression of INSM1 was found in all PanNET pure type cases. However, expression of INSM1 was negative in PDAC, ACC, and SPN in all cases, whereas faint expression was seen in the cytoplasm from SPN. MANEC comprises of two components: neuroendocrine carcinoma and adenocarcinoma components. The NET component was positive for INSM1 expression, whereas the PDAC component does not express INSM1, which aids in distinguishing these components. Our results suggest that INSM1 is a useful immunohistochemical marker for diagnosing pancreatic neuroendocrine tumor.

ARHGEF15 overexpression worsens the prognosis in patients with pancreatic ductal adenocarcinoma through enhancing the motility and proliferative activity of the cancer cells.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive neoplastic diseases, associated with a remarkably poor prognosis. However, the molecular mechanisms underlying the development of PDAC remain elusive. The aim of this study was to identify genes whose expressions are correlated with a poor prognosis in PDAC patients, and to unravel the mechanisms underlying the involvement of these genes in the development of the cancer. METHODS: Global gene expression profiling was conducted in 39 specimens obtained from Japanese patients with PDAC to identify genes whose expressions were correlated with a shorter overall survival. The effect of gene silencing or overexpression of ARHGEF15 in pancreatic cancer cell lines was examined by introducing siRNAs of ARHGEF15 or the ARHGEF15 expression vector. After assessing the effect of ARHGEF15 deregulation on the Rho-family proteins by pull-down assay, wound healing, transwell and cell viability assays were carried out to investigate the cellular phenotypes caused by the perturbation. RESULTS: The global mRNA expression profiling revealed that overexpression of ARHGEF15, a Rho-specific GEF, was significantly associated with a poor prognosis in patients with PDAC. We also found that the depletion of ARHGEF15 by RNA interference in pancreatic cancer cell lines downregulated the activities of molecules of the Rho signaling pathway, including RhoA, Cdc42 and Rac1. Then, we also showed that ARHGEF15 silencing significantly reduced the motility and viability of the cells, while its overexpression resulted in the development of the opposite phenotype in multiple pancreatic cancer cell lines. CONCLUSION: These data suggest that upregulation of ARHGEF15 contributes to the development of aggressive PDAC by increasing the growth and motility of the pancreatic cancer cells, thereby worsening the prognosis of these patients. Therefore, ARHGEF15 could serve as a novel therapeutic target in patients with PDAC.

Epithelial cell adhesion molecule-positive human hepatic neoplastic cells: development of combined hepatocellular-cholangiocarcinoma in mice.

BACKGROUND AND AIM: Human combined hepatocellular-cholangiocarcinoma (CHC) expresses several hepatic stem/progenitor cell (HSPC) markers, suggesting this neoplasm originates from HSPCs. We examined the significance of HSPC marker in CHC using a human CHC cell line. METHODS: We used a human CHC cell line (KMCH-1) previously established in our laboratory. The original tumor was classified as CHC, showing areas of typical hepatocellular carcinoma (HCC) and cholangiocarcinoma (ChC). We examined the expression of HSPC markers and hepatocyte markers in KMCH-1 by flow cytometry (FCM) and quantitative real-time polymerase chain reaction. EpCAM(+) and EpCAM(-) KMCH-1 cells were isolated. Subsequently, their morphological features, HSPC marker expression, and biological characteristics were examined in vitro and in vivo. RESULTS: FCM showed expression of EpCAM, K7, K19, and ABCG2 in KMCH-1, with various degrees. EpCAM(+) cells expressed K19 mRNA, but did not express α-fetoprotein (AFP). In contrast, EpCAM(-) cells expressed AFP mRNA, but did not express K19. EpCAM(+) cells produced both EpCAM(+) and EpCAM(-) cells, but EpCAM(-) cells produced only EpCAM(-) cells in vitro. EpCAM(+) cells showed higher tumorigenicity and formed larger tumors than EpCAM(-) cells. Inoculation of EpCAM(+) and EpCAM(-) cells produced both ChC and HCC-like component and HCC-like component only, respectively. CONCLUSION: It is speculated that some CHCs may originate from EpCAM(+) neoplastic cells, and that these cells may affect malignant behavior and progression in such CHCs.

Thermoresponsive nanostructured surfaces generated by the Langmuir-Schaefer method are suitable for cell sheet fabrication.

Thermoresponsive poly(N-isopropylacrylamide) (PIPAAm)-immobilized surfaces for controlling cell adhesion and detachment were fabricated by the Langmuir-Schaefer method. Block copolymers composed of polystyrene and PIPAAm (St-IPAAms) having various chain lengths and compositions were synthesized by reversible addition-fragmentation chain transfer radical polymerization. The St-IPAAm Langmuir film at an air-water interface was horizontally transferred onto a hydrophobically modified glass substrate while regulating its density. Atomic force microscopy images clearly visualized nanoscaled sea-island structures on the surface. By adjusting both the composition of St-IPAAms and the density of immobilized PIPAAms, a series of thermoresponsive surfaces was prepared to control the strength, rate, and quality of cell adhesion and detachment through changes in temperature across the lower critical solution temperature range of PIPAAm molecules. In addition, a two-dimensional cell structure (cell sheet) was more rapidly recovered on the optimized surfaces than on conventional PIPAAm surfaces. These unique PIPAAm surfaces are suggested to be useful for controlling the strength of cell adhesion and detachment.

Side population cell fractions from hepatocellular carcinoma cell lines increased with tumor dedifferentiation, but lack characteristic features of cancer stem cells.

BACKGROUND AND AIM: Cancer stem cells (CSCs), a minority population with stem cell-like characteristics, play important roles in cancer development and progression. Putative CSC markers, such as CD13, CD90, CD133, and epithelial cell adhesion molecule (EpCAM), and side population (SP) technique are generally used in an attempt to isolate CSCs. We aimed to clarify the relationship between CSCs and clonal dedifferentiation in hepatocellular carcinoma (HCC). METHODS: We used a well-differentiated HCC cell line (HAK-1A) and a poorly differentiated HCC cell line (HAK-1B) established from a single nodule with histological heterogeneity. HAK-1B arose because of clonal dedifferentiation of HAK-1A. The SP cells and non-SP (NSP) cells were isolated from the two cell lines with a FACSAria II and used for the analyses. RESULTS: The SP cell fractions in HAK-1A and HAK-1B were 0.2% and 0.9%, respectively. CD90 or EpCAM was not expressed in either HAK-1A or HAK-1B, while CD13 and CD133 were expressed in HAK-1B alone. Although sphere forming ability, tumorigenicity, growth rate, and CD13 expression were higher in HAK-1B SP cells than HAK-1B NSP cells, there were no differences in drug resistance, colony forming ability, or cell cycle rates between HAK-1B SP and NSP cells, suggesting HAK-1B SP cells do not fulfill CSC criteria. CONCLUSIONS: Our findings suggested a possible relationship between the expression of CSC markers and clonal dedifferentiation. However, the complete features of CSC could not be identified in SP cells, and the concept of SP cells as a universal marker for CSC may not apply to HAK-1A and HAK-1B.

Evaluation of endoscopic transpapillary brushing cytology for the diagnosis of bile duct cancer based on the histopathologic findings.

BACKGROUND AND AIM: Diagnosis of the bile duct cancer still needs more accuracy. Studies on endoscopic retrograde cholangiopancreatography (ERCP)-guided brushing cytology were carried to evaluate the role of the endoscopic transpapillary brushing cytology for the diagnosis of bile duct cancer. PATIENTS AND METHOD: The study involved 76 consecutive patients who underwent ERCP-guided bile duct cytology for the diagnosis of bile duct cancer from 2008 to August 2012. Three types of cytological specimens were obtained using different sampling methods, i.e., bile aspiration cytology (BAC), brush tip cytology (BTC), and post brushing bile cytology (PBC), to investigate their diagnostic abilities, and comparatively studied with each macroscopic type of the surgically resected specimens. RESULTS: The cancer-positive rate was 67.1 % (BAC alone: 41.9 %), and the use of BTC and PBC in addition to BAC yielded a statistically significant increase of the cancer-positive rate (p = 0.0031). In 34 resected cases, the cancer-positive rate in relation to the macroscopic type was improved by the addition of BTC and PBC to BAC alone for the papillary (87.5 vs. 40.0 %, p = 0.071) and nodular (100 vs. 70.0 %, p = 0.0603) types, but not for the flat type (62.5 vs. 57.1 %; p = 0.7651). CONCLUSION: The diagnostic ability of ERCP-guided brushing cytology could be improved by the addition of PBC. However, the cancer-positive rate was the lowest for the flat type of bile duct cancer.

Hyperthermia in the treatment of recurrent abdominal desmoid tumor.

The most suitable management of recurrent abdominal desmoid tumor is still unclear. A case of recurrent huge abdominal desmoid tumor successfully treated by hyperthermia therapy is described. A 63-year-old man was operated upon for desmoid tumor in the retroperitoneum involving pancreas, posterior wall of the stomach and transverse mesocolon in 2007. In 2008, the tumor recurred and could not be resected because of the patient refused the operation. Several therapies using tamoxifen, anastrozole, imatinib mesylate and radiotherapy were all ineffective. The tumor grew bigger and bigger during a treatment period. Finally, hyperthermia treatment was applied to the tumor. The size of the recurrent desmoid tumor reduced 75% by hyperthermia treatment for 46-month. Base on this experience, we recommend hyperthermia as the treatment for patients with recurrent abdominal desmoid tumor that several therapeutic strategies did not achieve a remarkable response.

Immunophenotypes and Tumor Immune Microenvironment in Hepatocellular Carcinoma With Macrotrabecular Massive and Vessels Encapsulating Tumor Clusters.

BACKGROUND/AIM: Recently, vessels encapsulating tumor clusters (VETC) pattern and macrotrabecular massive (MTM) pattern of hepatocellular carcinoma (HCC) have been reported as aggressive histological types. These histological patterns showed an immunosuppressive tumor immune microenvironment (TIME). Since there have been no reports on the differences of these two subtypes simultaneously, this study examined the immunophenotypes and TIME of MTM-HCC and VETC-HCC immunohistochemically. PATIENTS AND METHODS: Seventy-four cases of previously diagnosed HCC, including 32 MTM-HCCs, 21 VETC-HCCs, and 21 conventional HCCs, were enrolled in immunohistochemical analysis. We conducted immunohistochemical analysis. RESULTS: We found that MTM-HCC showed less frequent expression of HepPar-1, which is one of the most common hepatocytic markers. In MTM-HCC, the frequency of high expression levels of Keratin19, carbonic anhydrase (CA) IX, and PD-L1 was higher compared to VETC-HCC and conventional HCC. PD-L1 expression was found in 34.4% of MTM-HCC, 0% of VETC-HCC, and 19.0% of conventional HCC. The rate of PD-L1 expression in MTM-HCC was significantly higher than the others (p=0.0015). PD-L1 expression was significantly associated with epithelial cell adhesion molecules and CA IX expression, which are representative markers of tumor stemness and hypoxic conditions, respectively. The CD8 infiltration in VETC-HCC was significantly lower than that in conventional HCC. CONCLUSION: MTM-HCC had different immunophenotypes and TIMEs compared to HCC with the VETC pattern. Although both had immunosuppressive TIME, the elements forming TIME were quite different. To enhance the immune checkpoint inhibitor efficacy, changing TIME from a suppressive to an active form is essential.

Terminal-functionality effect of poly(N-isopropylacrylamide) brush surfaces on temperature-controlled cell adhesion/detachment.

Terminally functionalized poly(N-isopropylacrylamide) (PIPAAm) brush grafted glass surfaces were prepared by a surface-initiated reversible addition-fragmentation chain transfer radical (SI-RAFT) polymerization. SI-RAFT mediated PIPAAm chains possessed terminal dodecyl trithiocarbonate groups which can be substituted with various functional groups. In this study, dodecyl groups were substituted with hydrophilic maleimide groups for controlling the thermoresponsive character of PIPAAm brushes. PIPAAm brushes exhibited reversible temperature-dependent surface wettability changes around PIPAAm's lower critical solution temperature. Phase transition of dodecyl-terminated PIPAAm brushes clearly shifted to lower temperature than that of maleimide-terminated PIPAAm brushes, and this shift was attributed to promoted PIPAAm dehydration via terminal hydrophobes. By using this feature, the specific adhesion temperatures of bovine carotid artery endothelial cells (BAECs) on the PIPAAm brush surfaces were successfully controlled. BAECs were initiated to adhere on dodecyl-PIPAAm surfaces at 31 °C, while their adhesion was significantly suppressed on maleimide-PIPAAm surfaces under 33 °C. In contrast, terminal functionality scarcely affected the thermoresponsive behavior of PIPAAm brushes in the polymer rehydration process by reducing temperatures, and thus, the difference in spontaneous cell detachment from different PIPAAm-brush surface was negligible. Consequently, confluently cultured cells were able to be harvested as contiguous cell sheets from individual surfaces with comparable periods at 20 °C.

Lymph node metastatic status could predict the prognosis of intracholecystic papillary neoplasm of gallbladder.

Intracholecystic papillary neoplasm (ICPN) is a non-invasive epithelial tumor that presents as a grossly identifiable mass arising in the mucosa and protruding into the lumen. ICPN is associated with invasive carcinoma. There are few studies on the clinicopathological features of ICPN, including that with invasive carcinoma. We evaluated the clinicopathological characteristics of 42 ICPNs and 41 conventional gallbladder adenocarcinomas (cGBAs). Subserosa or deeper (≥ss) invasion was significantly lower in ICPN (61.9%) than that in cGBA (90.2%) (P = 0.004). Cox regression analysis revealed that lymph node metastasis (hazard ratio [HR] [95% confidence interval (CI)]: 2.610 [1.131, 6.024], P = 0.025) and positive margin (HR [95% CI]: 5.143 [2.113, 12.516], P < 0.001), but not ≥ss invasion (HR [95% CI]: 1.541 [0.479, 4.959], P = 0.469), were independent prognostic factors. In addition, there was a significant interaction between histological type and lymph node metastasis (HR [95% CI]: 0.191 [0.042, 0.983], P = 0.033). In cGBA, the presence or absence of lymph node metastasis did not affect prognosis; however, ICPN without lymph node metastasis had better prognosis. Therefore, the histological classification of ICPN and cGBA and the pathological evaluation of lymph node metastasis in ICPN are crucial for determining prognosis.

Clinicopathological characteristics and molecular analysis of lymphocyte-rich hepatocellular carcinoma.

Lymphocyte-rich hepatocellular carcinoma (LR-HCC), a newly proposed subtype of HCC, is characterized with abundant lymphocyte infiltration in the tumor. LR-HCC has a relatively good prognosis and is quite rare (<1% of all HCC). We examined LR-HCC clinicopathological and molecular characteristics by analyzing 451 surgically resected HCC cases without any prior treatment history at our hospital between 2012 and 2021. Clinicopathological features of LR-HCC and other HCCs (non-LR-HCC) were compared. Neoplastic and nonneoplastic hepatocytes from LR-HCC (n = 4) were collected with a laser microdissection system; RNA was extracted, followed by microarray analysis to examine lymphocytic infiltration-related molecular targets. Immunohistochemical staining of identified molecular target was performed in LR-HCC and non-LR-HCC. CD3, CD20, and CD8 immunostaining was also performed in LR-HCCs. There were 28 cases of LR-HCC (6%). No statistically significant differences were found in clinicopathological features, except for gross type, between LR-HCC and non-LR-HCC cases. The LR-HCC 5-year survival rate was >90%. Microarray analysis revealed high CCL20 expression in LR-HCC cases; immunohistochemical study showed significantly higher CCL20 expression in LR-HCC (P < 0.01) than in non-LR-HCC. CCR6, the only CCL20 receptor, was observed in infiltrating lymphocytes and HCC cells in LR-HCC. There were significantly more CD3-positive cells than CD20-positive cells (P < 0.0001) in tumor-infiltrating lymphocytes, most of which were CD8-positive T cells. In conclusion, there were no significant differences in clinicopathological characteristics between LR-HCC and non-LR-HCC, except for gross and LR microscopic features. CCL20 expression in LR-HCC may contribute to infiltration of large numbers of CD8-positive lymphocytes.

Liposarcoma of the pleural cavity.

Liposarcoma rarely occurs in the pleura or thoracic cavity, and few reports appear in the literature. We hypothesized that combining clinicopathologic, immunohistochemical, and fluorescence in situ hybridization methods would allow definite diagnoses. Using formalin-fixed, paraffin-embedded blocks, we examined 6 atypical lipomatous tumor/well-differentiated liposarcomas (ALT/WDLPS), 5 dedifferentiated liposarcomas (DDLPSs), 2 pleomorphic liposarcomas, and 1 myxoid liposarcoma (MLPS). We used the Kaplan-Meier method and the Wilcoxon test for survival analysis for prognostic factor evaluation. Histologically, ALT/WDLPS was composed of a relatively mature adipocytic proliferation, accompanied by some lipoblasts. DDLPS exhibited round-to-oval tumor cells with a high nucleus-to-cytoplasm ratio that had proliferated in nests, accompanied in case 10 by some giant cells but no fatty cells. The pleomorphic type contained a varying proportion of pleomorphic lipoblasts. MLPS displayed uniform round- to oval-shaped cells and small signet-ring lipoblasts in a myxoid stroma. Immunohistochemically, 11 (79%), 11 (79%), and 10 (71%) of 14 cases were positive for S-100, p16, and CDK4, respectively. Six of the 14 cases (43%) were positive for MDM2 and adipophilin. One case of ALT/WDLPS and 3 cases of DDLPS exhibited MDM2 amplification by fluorescence in situ hybridization (Vysis LSI MDM2 SpectrumGreen Probe plus Vysis CEP 12 SpectrumOrange probe). ALT/WDLPS was the most favorable type for survival, while adipophilin tended to be a negative prognostic factor for pleural liposarcoma. For a firm diagnosis of liposarcoma in the pleura, immunohistochemistry for CDK4, MDM2, and adipophilin together with MDM2 gene amplification by fluorescence in situ hybridization may be an important diagnostic tool.

Tumor-derived insulin-like growth factor-binding protein-1 contributes to resistance of hepatocellular carcinoma to tyrosine kinase inhibitors.

BACKGROUND: Antiangiogenic tyrosine kinase inhibitors (TKIs) provide one of the few therapeutic options for effective treatment of hepatocellular carcinoma (HCC). However, patients with HCC often develop resistance toward antiangiogenic TKIs, and the underlying mechanisms are not understood. The aim of this study was to determine the mechanisms underlying antiangiogenic TKI resistance in HCC. METHODS: We used an unbiased proteomic approach to define proteins that were responsible for the resistance to antiangiogenic TKIs in HCC patients. We evaluated the prognosis, therapeutic response, and serum insulin-like growth factor-binding protein-1 (IGFBP-1) levels of 31 lenvatinib-treated HCC patients. Based on the array of results, a retrospective clinical study and preclinical experiments using mouse and human hepatoma cells were conducted. Additionally, in vivo genetic and pharmacological gain- and loss-of-function experiments were performed. RESULTS: In the patient cohort, IGFBP-1 was identified as the signaling molecule with the highest expression that was inversely associated with overall survival. Mechanistically, antiangiogenic TKI treatment markedly elevated tumor IGFBP-1 levels via the hypoxia-hypoxia inducible factor signaling. IGFBP-1 stimulated angiogenesis through activation of the integrin α5ß1-focal adhesion kinase pathway. Consequently, loss of IGFBP-1 and integrin α5ß1 by genetic and pharmacological approaches re-sensitized HCC to lenvatinib treatment. CONCLUSIONS: Together, our data shed light on mechanisms underlying acquired resistance of HCC to antiangiogenic TKIs. Antiangiogenic TKIs induced an increase of tumor IGFBP-1, which promoted angiogenesis through activating the IGFBP-1-integrin α5ß1 pathway. These data bolster the application of a new therapeutic concept by combining antiangiogenic TKIs with IGFBP-1 inhibitors.

Terminally functionalized thermoresponsive polymer brushes for simultaneously promoting cell adhesion and cell sheet harvest.

For preparing cell sheets effectively for cell sheet-based regenerative medicine, cell-adhesion strength to thermoresponsive cell culture surfaces need to be controlled precisely. To design new thermoresponsive surfaces via a terminal modification method, thermoresponsive polymer brush surfaces were fabricated through the surface-initiated reversible addition-fragmentation chain transfer (RAFT) radical polymerization of N-isopropylacrylamide (IPAAm) on glass substrates. The RAFT-mediated grafting method gave dithiobenzoate (DTB) groups to grafted PIPAAm termini, which can be converted to various functional groups. In this study, the terminal carboxylation of PIPAAm chains provided high cell adhesive property to thermoresponsive surfaces. Although cell adhesion is generally promoted by a decrease in the grafted PIPAAm amount, the decrease also decelerated thermally-induced cell detachment, whereas the influence of terminal modification was negligible on the cell detachment. Consequently, the terminally modified PIPAAm brush surfaces allowed smooth muscle cells (SMCs) to simultaneously adhere strongly and detach themselves rapidly. In this study, SMCs were unable to reach a confluent monolayer on as-prepared PIPAAm brush surfaces (grafted amount: 0.41 µg/cm(2)) without terminal carboxylation due to their insufficient cell-adhesion strength. On the other hand, though a decrease in the PIPAAm amount allowed SMCs to form a confluent cell monolayer on the PIPAAm brush surface, the SMCs were unable to be harvested as a monolithic cell sheet by low-temperature culture at 20 °C. Because of their unique property, only terminal-carboxylated PIPAAm brush surfaces achieved rapid harvesting of complete cell sheets by low-temperature culturing.

Metastatic pulmonary adenocarcinoma 13 years after curative resection for pancreatic cancer: report of a case and review of Japanese literature.

CONTEXT: For the majority of patients, ductal adenocarcinoma of the pancreas remains a lethal disease. Currently, surgical extirpation for localized disease offers the only chance for long-term survival. CASE REPORT: We report a patient who underwent successful resection of isolated lung metastasis occurring 13 years after pancreatic cancer resection. A 59-year-old woman underwent distal pancreatectomy for pancreatic cancer 13 years previously, followed by adjuvant chemotherapy, and was followed-up at the outpatient clinic of a local hospital. From around June 2010, she noticed bloody sputum, so she visited a local hospital. Since her chest X-ray and CT revealed a 1.5 cm mass shadow in the segment 10 of her right lung and she was referred to the Respiratory Disease Center of our hospital. As a result of through examinations, she was strongly suspected of having lung metastasis of pancreatic cancer, and underwent partial pneumonectomy. Postoperative histopathological examination of the resected specimen was consistent with lung metastasis of pancreatic cancer. She is still alive and currently receives third line of chemotherapy. CONCLUSION: Patients who have achieved long-term survival after pancreatic cancer resection and can tolerate surgery may benefit from resection of a lung metastasis of pancreatic cancer in terms of survival, if it controls the metastasis.