Claudio De Vito

Transportin Regulates Nuclear Import of CD44

CD44 is a facultative cell surface proteoglycan that serves as the principal cell surface receptor for hyaluronan (HA). Studies have shown that in addition to participating in numerous signaling pathways, CD44 becomes internalized upon engagement by ligand and that a portion of its intracellular domain can translocate to the nucleus where it is believed to play a functional role in cell proliferation and survival. However, the mechanisms whereby fragments of CD44 enter the nucleus have not been elucidated. Here we show that CD44 interacts with two import receptors of the importin β superfamily, importin β itself and transportin. Inhibition of importin β-dependent transport failed to block CD44 accumulation in the nucleus. By contrast, inhibition of the transportin-dependent pathway abrogated CD44 import. Mutagenesis of the intracellular domain of CD44 revealed that the 20 membrane-proximal residues contain sequences required for transportin-mediated nuclear transport. Our observations provide evidence that CD44 interacts with importin family members and identify the transportin-dependent pathway as the mechanism whereby full-length CD44 enters the nucleus.

Pre-retrieval reperfusion decreases cancer recurrence after rat ischemic liver graft transplantation

BACKGROUND & AIMS Liver transplantation from marginal donors is associated with ischemia/reperfusion (I/R) lesions, which may increase the risk of post-transplant hepatocellular carcinoma (HCC) recurrence. Graft reperfusion prior to retrieval (as for extracorporeal membrane oxygenation--ECMO) can prevent I/R lesions. The impact of I/R on the risk of cancer recurrence was assessed on a syngeneic Fischer-rat liver transplantation model. METHODS HCC cells were injected into the vena porta of all recipients at the end of an orthotopic liver transplantation (OLT). Control donors were standard heart-beating, ischemic ones (ISC), underwent 10 min or 30 min inflow liver clamping prior to retrieval, and ischemic/reperfused (ISC/R) donors underwent 2h liver reperfusion after the clamping. RESULTS I/R lesions were confirmed in the ISC group, with the presence of endothelial and hepatocyte injury, and increased liver function tests. These lesions were in part reversed by the 2h reperfusion in the ISC/R group. HCC growth was higher in the 10 min and 30 min ISC recipients (p = 0.018 and 0.004 vs. control, as assessed by MRI difference between weeks one and two), and was prevented in the ISC/Rs (p = 0.04 and 0.01 vs. ISC). These observations were associated with a stronger pro-inflammatory cytokine profile in the ISC recipients only, and the expression of hypoxia and HCC growth-enhancer genes, including Hmox1, Hif1a and Serpine1. CONCLUSIONS This experiment suggests that ischemia/reperfusion lesions lead to an increased risk of post-transplant HCC recurrence and growth. This observation can be reversed by graft reperfusion prior to retrieval.

Histological heterogeneity in primary and metastatic classic combined hepatocellular-cholangiocarcinoma: a case series

Combined hepatocellular-cholangiocarcinoma (cHCC-CC) is a rare and aggressive primary liver cancer with both hepatocellular and cholangiocellular differentiation. Due to its bi-phenotypic component, cHCC-CC is a heterogeneous tumour and histopathological analysis of metastatic deposits is poorly characterized. In this retrospective study, we describe four patients in whom the histology from resected specimens of both primary and recurrent and/or metastatic tumour was available for comparison and immunohistochemical characterization. Our study shows that recurrent or metastatic deposits replicate the heterogeneity of the primary cHCC-CC, that even originally small foci of divergent differentiation can become predominant later on and that hepatocellular and cholangiocellular components can show different tropism in distant organs. In our experience, the behaviour of recurrent/metastatic cHCC-CC is unpredictable and histological examination is necessary to guide treatment options at present.

Unexpected Steatotic Liver Lesions

Gastroenterology 2016;151:32–33 Question: A 22year-old woman with no past medical history was investigated for hypoglycemia episodes. A nodule located in the head of the pancreas was identified, with radiologic features of a neuroendocrine neoplasm. The overall clinical presentation was consistent with an insulinoma. No distant lesionwas detected. She underwent a Whipple’s procedure, and the histopathological examination reported a 2.2-cm, welldifferentiated neuroendocrine tumor (insulinoma) G2 (4% Ki-67 index), with no lymphovascular invasion or lymph nodes metastasis (0 of 30 lymph nodes). Six months later, the patient was asymptomatic, and a follow-up (CT) scan, completed by magnetic resonance imaging and dihydroxyphenylalanine positron emission tomography–CT, revealed multiple bilobar steatotic liver areas, but no sign of metastasis (Figure A, B). No hypermetabolic activity was identified by positron emission tomography. A CT-guided biopsy of 1 lesion showed steatotic hepatocytes with a loss of liver fatty acid-binding protein (L-FABP), and no evidence for neuroendocrine tumor metastasis (Figure C [stain: hematoxylin and eosin] and D [L-FABP immunostaining]). What is the diagnosis? Look on page 33 for the answer and see the Gastroenterology website (www.gastrojournal.org) for more information on submitting your favorite image to Clinical Challenges and images in GI.

Histopathology of livers in patients with congenital portosystemic shunts (Abernethy malformation): a case series of 22 patients

Congenital portosystemic shunt (CPSS) is a congenital anomaly resulting in partial or complete diversion of the portal blood into the systemic circulation. The literature on the histological changes in livers of patients with CPSS is limited. Liver histology of 22 consecutive patients managed in our institution between 2001 and 2016 was reviewed. Twenty-one patients were children at the time of diagnosis. Thirty-two specimens were available and consisted of three explant livers and 29 biopsy samples from 19 patients. Sixteen samples were from wedge biopsies taken at the time of shunt closure. Thirteen were from core needle biopsies taken during clinical work-up. A variable proportion of portal tracts contained prominent thin-walled channels (PTWCs) and arterio-biliary dyads. The proportion of portal tracts containing triads, arterio-biliary dyads and biliary monads varied considerably in the different samples. Dilated inlet venules, increase in the number of portal arteries or the presence of portal arteries of increased size, deposition of copper-associated protein, sinusoidal dilatation, capillarization and intralobular individual arteries were present. Physiological nuclear vacuolation of periportal hepatocytes was absent in most samples from our paediatric patients. Presence of PTWCs, arterial-biliary dyads, increased arterial profiles in portal tracts and lobule and lack of the physiological periportal vacuolated hepatocytes in children are the most characteristic histological changes of CPSS in the liver periphery.

Identification of Differential Transcriptional Patterns in Primary and Secondary Hyperparathyroidism

Context Hyperparathyroidism is associated with hypercalcemia and the excess of parathyroid hormone secretion; however, the alterations in molecular pattern of functional genes during parathyroid tumorigenesis have not been unraveled. We aimed at establishing transcriptional patterns of normal and pathological parathyroid glands (PGs) in sporadic primary (HPT1) and secondary hyperparathyroidism (HPT2). Objective To evaluate dynamic alterations in molecular patterns as a function of the type of PG pathology, a comparative transcript analysis was conducted in subgroups of healthy samples, sporadic HPT1 adenoma and hyperplasia, and HPT2. Design Normal, adenomatous, HPT1, and HPT2 hyperplastic PG formalin-fixed paraffin-embedded samples were subjected to NanoString analysis. In silico microRNA (miRNA) analyses and messenger RNA-miRNA network in PG pathologies were conducted. Individual messenger RNA and miRNA levels were assessed in snap-frozen PG samples. Results The expression levels of c-MET, MYC, TIMP1, and clock genes NFIL3 and PER1 were significantly altered in HPT1 adenoma compared with normal PG tissue when assessed by NanoString and quantitative reverse transcription polymerase chain reaction. RET was affected in HPT1 hyperplasia, whereas CaSR and VDR transcripts were downregulated in HPT2 hyperplastic PG tissue. CDH1, c-MET, MYC, and CaSR were altered in adenoma compared with hyperplasia. Correlation analyses suggest that c-MET, MYC, and NFIL3 exhibit collective expression level changes associated with HPT1 adenoma development. miRNAs, predicted in silico to target these genes, did not exhibit a clear tendency upon experimental validation. Conclusions The presented gene expression analysis provides a differential molecular characterization of PG adenoma and hyperplasia pathologies, advancing our understanding of their etiology.

Deficient natural killer cell NKp30‐mediated function and altered NCR3 splice variants in hepatocellular carcinoma

The activating natural cytotoxicity receptor NKp30 is critical for natural killer (NK) cell function and tumor immune surveillance. The natural cytotoxicity receptor‐3 (NCR3) gene is transcribed into several splice variants whose physiological relevance is still incompletely understood. In this study, we investigated the role of NKp30 and its major ligand B7 homolog 6 (B7‐H6) in patients with hepatocellular carcinoma (HCC). Peripheral blood NK cell phenotype was skewed toward a defective/exhausted immune profile with decreased frequencies of cells expressing NKp30 and natural killer group 2, member D and an increased proportion of cells expressing T‐cell immunoglobulin and mucin‐domain containing‐3. Moreover, NKp30‐positive NK cells had a reduced expression of NCR3 immunostimulatory splice variants and an increased expression of the inhibitory variant in patients with advanced tumor, resulting in deficient NKp30‐mediated functionality. Tumor‐infiltrating lymphocytes showed a prevalent inhibitory NKp30 isoform profile, consistent with decreased NKp30‐mediated function. Of note, there were significant differences in the cytokine milieu between the neoplastic and the surrounding non‐neoplastic tissue, which may have further influenced NKp30 function. Exposure of NK cells to B7‐H6‐expressing HCC cells significantly down‐modulated NKp30, that was prevented by small interfering RNA–mediated knockdown, suggesting a role for this ligand in inhibiting NKp30‐mediated responses. Interestingly, B7‐H6 expression was reduced in HCC tissue and simultaneously augmented as a soluble form in HCC patients, particularly those with advanced staging or larger nodule size. Conclusion: These findings provide evidence in support of a role of NKp30 and its major ligand in HCC development and evolution.

Utility of MRI with morphologic and diffusion weighted imaging in the detection of post-treatment nodal disease in head and neck squamous cell carcinoma

PURPOSE To determine the diagnostic performance of morphologic MRI with diffusion weighted imaging (DWIMRI) for the detection of post-treatment lymph node (LN) recurrence of head and neck squamous cell carcinoma (HNSCC). METHODS This retrospective study is based on 33 HNSCC patients who underwent DWIMRI with apparent diffusion coefficient (ADC) measurements for suspected post-treatment loco-regional failure. Two radiologists, blinded to clinical/histopathological data, analyzed MR images according to established morphologic criteria and measured ADC values by drawing regions of interest on each normal/abnormal looking lymph node (LN). Histopathological findings in 40 neck dissections, 133 LN-levels and 755 LNs served as gold standard. RESULTS Malignant LNs had lower ADCmean values than benign LNs (1.15 ± 0.35 × 10-3 mm2/s versus 1.28 ± 0.28 × 10-3 mm2/s, p = .028). The optimal ADCmean threshold to differentiate malignant from benign LNs was 1.1695 × 10-3 mm2/s. Sensitivity, specificity, positive (PPV) and negative (NPV) predictive values (95%CI in parentheses) of DWIMRI with morphologic criteria and ADCmean <1.1695 × 10-3 mm2/s were: (a) 100%(86.2;100), 44.4%(15.3;77.3), 86.1%(69.7;94.7), and 100%(39.5;100) per neck dissection; (b) 83.6%(69.7;92.2), 91.6%(83.0;96.2), 85.4%(71.6;93.4), and 90.5%(81.7;95.5) per LN-level; (c) 53.1%(43.5;62.4), 95.5%(93.5;96.9), 67.4%(56.6;76.7), and 92.0%(89.6;93.9) per LN, respectively. CONCLUSION The high NPV of DWIMRI irrespective of analysis type (per neck dissection/per neck level/per lymph node) make it a useful follow-up tool after treatment.

Macrofollicular Variant of Follicular Thyroid Carcinoma: A Case Report

A 37-year-old female known for systemic lupus erythematosus underwent a right thyroid lobectomy for a growing nodule in the right lobe. A fine-needle aspiration biopsy (FNAB) performed 4 years before surgery reported a microfollicular proliferation. A second FNAB performed a few months before surgery revealed a benign lesion. The surgical specimen was fixed in 4 % buffered formaldehyde. The entire nodule was sampled, with particular attention to the capsule, and embedded in paraffin and corresponding sections were stained with hematoxylin and eosin for histological examination. Immunohistochemistry using HBME1 and galectin-3 was performed.