Recurrent loss of heterozygosity correlates with clinical outcome in pancreatic neuroendocrine cancer.

Pancreatic neuroendocrine tumors (pNETs) are uncommon cancers arising from pancreatic islet cells. Here we report the analysis of gene mutation, copy number, and RNA expression of 57 sporadic well-differentiated pNETs. pNET genomes are dominated by aneuploidy, leading to concordant changes in RNA expression at the level of whole chromosomes and chromosome segments. We observed two distinct patterns of somatic pNET aneuploidy that are associated with tumor pathology and patient prognosis. Approximately 26% of the patients in this series had pNETs with genomes characterized by recurrent loss of heterozygosity (LoH) of 10 specific chromosomes, accompanied by bi-allelic MEN1 inactivation and generally poor clinical outcome. Another ~40% of patients had pNETs that lacked this recurrent LoH pattern but had chromosome 11 LoH, bi-allelic MEN1 inactivation, and universally good clinical outcome. The somatic aneuploidy allowed pathogenic germline variants (e.g., ATM) to be expressed unopposed, with RNA expression patterns showing inactivation of downstream tumor suppressor pathways. No prognostic associations were found with tumor morphology, single gene mutation, or expression of RNAs reflecting the activity of immune, differentiation, proliferative or tumor suppressor pathways. In pNETs, single gene mutations appear to be less important than aneuploidy, with MEN1 the only statistically significant recurrently mutated driver gene. In addition, only one pNET in the series had clearly actionable single nucleotide variants (SNVs) (in PTEN and FLCN) confirmed by corroborating RNA expression changes. The two clinically relevant patterns of LoH described here define a novel oncogenic mechanism and a plausible route to genomic precision oncology for this tumor type.

Profound Reversible Hypogammaglobulinemia Caused by Celiac Disease in the Absence of Protein Losing Enteropathy.

When patients with hypogammaglobulinemia are encountered, a vigorous search should be undertaken for secondary treatable causes. Here we describe the first case of a patient with severe asymptomatic hypogammaglobulinemia where the underlying cause was undiagnosed celiac disease. A strict gluten free diet resulted in resolution of her mild long-standing abdominal symptoms and correction of her hypogammaglobulinemia. There was corresponding improvement in her duodenal histology and normalisation of her celiac serology. Protein losing enteropathy was unlikely to have been the mechanism of her profound hypogammaglobulinemia, as her albumin was within the normal range and she had a normal fecal alpha 1 antitrypsin level. Application of the Ameratunga et al. (2013) diagnostic criteria was helpful in confirming this patient did not have Common Variable Immunodeficiency Disorder (CVID). Celiac disease must now be considered in the differential diagnosis of severe hypogammaglobulinemia. There should be a low threshold for undertaking celiac serology in patients with hypogammaglobulinemia, even if they have minimal symptoms attributable to gut disease.

Malignant granular cell tumour of the mediastinum.

This report describes a case of malignant granular cell tumour arising in the mediastinum, detailing the investigations undertaken to reach this rare diagnosis. A 63-year-old man was referred from the Pacific Islands for investigation of a 8cm mediastinal mass extending into the left pleura and associated with pleural nodules and pleural effusion. Needle aspiration via bronchoscopy yielded insufficient material for cytological interpretation and needle biopsy showed normal respiratory epithelium. CT-guided FNA revealed scattered large polygonal to spindle cells with granular cytoplasm and indistinct borders. The needle core biopsy yielded scanty cells with abundant granular cytoplasm, oval and regular nuclei which were moderately positive for CD68, vimentin and S100 and negative for CKMNF116, CK5/6, CK7, CK20, TTF-1, chromogranin and synaptophysin. In view of the benign morphology, these cells were interpreted to be histiocytes. The incisional biopsy revealed cords and trabeculae of cells identical to the CT samples. These cells were polygonal with abundant granular cytoplasm. Some cells showed large eosinophilic cytoplasmic globules not seen in the FNA sample. The tumour was however, heterogeneous in appearance with some areas exhibiting criteria of malignancy: necrosis, vesicular nuclei with large nucleoli, high nuclear-to-cytoplasmic ratio and nuclei pleomorphism. In addition, p53 expression in 10% of tumour nuclei, a high Ki67 proliferative rate (>10%), the deep seated location and extension of the tumour into adjacent organs favoured a diagnosis of malignancy.

Ob/ob mouse livers show decreased oxidative phosphorylation efficiencies and anaerobic capacities after cold ischemia.

BACKGROUND: Hepatic steatosis is a major risk factor for graft failure in liver transplantation. Hepatic steatosis shows a greater negative influence on graft function following prolonged cold ischaemia. As the impact of steatosis on hepatocyte metabolism during extended cold ischaemia is not well-described, we compared markers of metabolic capacity and mitochondrial function in steatotic and lean livers following clinically relevant durations of cold preservation. METHODS: Livers from 10-week old leptin-deficient obese (ob/ob, n = 9) and lean C57 mice (n = 9) were preserved in ice-cold University of Wisconsin solution. Liver mitochondrial function was then assessed using high resolution respirometry after 1.5, 3, 5, 8, 12, 16 and 24 hours of storage. Metabolic marker enzymes for anaerobiosis and mitochondrial mass were also measured in conjunction with non-bicarbonate tissue pH buffering capacity. RESULTS: Ob/ob and lean mice livers showed severe (>60%) macrovesicular and mild (<30%) microvesicular steatosis on Oil Red O staining, respectively. Ob/ob livers had lower baseline enzymatic complex I activity but similar adenosine triphosphate (ATP) levels compared to lean livers. During cold storage, the respiratory control ratio and complex I-fueled phosphorylation deteriorated approximately twice as fast in ob/ob livers compared to lean livers. Ob/ob livers also demonstrated decreased ATP production capacities at all time-points analyzed compared to lean livers. Ob/ob liver baseline lactate dehydrogenase activities and intrinsic non-bicarbonate buffering capacities were depressed by 60% and 40%, respectively compared to lean livers. CONCLUSIONS: Steatotic livers have impaired baseline aerobic and anaerobic capacities compared to lean livers, and mitochondrial function indices decrease particularly from after 5 hours of cold preservation. These data provide a mechanistic basis for the clinical recommendation of shorter cold storage durations in steatotic donor livers.

Inflammatory pseudotumor-like follicular dendritic cell tumor of liver and spleen: granulomatous and eosinophil-rich variants mimicking inflammatory or infective lesions.

Inflammatory pseudotumor-like follicular dendritic cell (FDC) tumor is an uncommon, Epstein-Barr virus-associated neoplasm of the liver or spleen, characterized by spindly tumor cells dispersed in a background of small lymphocytes and plasma cells. We report 6 diagnostically challenging cases in which the neoplastic component is further overshadowed by granulomas or eosinophils. The patients included 2 men and 4 women with a median age of 45.5 years, and 1 of them showed concurrent involvement of the liver and spleen. The presence of extensive coalescent epithelioid granulomas in 3 splenic tumors and 3 liver tumors raised the possibilities of an infective process or sarcoidosis. In another liver tumor, the massive infiltrate of eosinophils, accompanied by geographic eosinophilic abscesses, suggested parasitic infestation or so-called eosinophilic granuloma of the liver. However, scrutiny of the tissue between the granulomas or among the eosinophils revealed scattered atypical spindly cells with indistinct cell borders, large vesicular nuclei, and distinct nucleoli. The atypical cells were positive for FDC markers on immunostaining (CD21, CD35) and Epstein-Barr virus on in situ hybridization. Thus, a diagnosis of inflammatory pseudotumor-like FDC tumor could be confirmed. Awareness of the granulomatous and eosinophil-rich variants of this tumor type will facilitate the correct diagnosis to be made.

Malignant granular cell tumour of the mediastinum

This report describes a case of malignant granular cell tumour arising in the mediastinum, detailing the investigations undertaken to reach this rare diagnosis. A 63-year-old man was referred from the Pacific Islands for investigation of a 8cm mediastinal mass extending into the left pleura and associated with pleural nodules and pleural effusion. Needle aspiration via bronchoscopy yielded insufficient material for cytological interpretation and needle biopsy showed normal respiratory epithelium. CT-guided FNA revealed scattered large polygonal to spindle cells with granular cytoplasm and indistinct borders. The needle core biopsy yielded scanty cells with abundant granular cytoplasm, oval and regular nuclei which were moderately positive for CD68, vimentin and S100 and negative for CKMNF116, CK5/6, CK7, CK20, TTF-1, chromogranin and synaptophysin. In view of the benign morphology, these cells were interpreted to be histiocytes. The incisional biopsy revealed cords and trabeculae of cells identical to the CT samples. These cells were polygonal with abundant granular cytoplasm. Some cells showed large eosinophilic cytoplasmic globules not seen in the FNA sample. The tumour was however, heterogeneous in appearance with some areas exhibiting criteria of malignancy: necrosis, vesicular nuclei with large nucleoli, high nuclear-to-cytoplasmic ratio and nuclei pleomorphism. In addition, p53 expression in 10% of tumour nuclei, a high Ki67 proliferative rate (>10%), the deep seated location and extension of the tumour into adjacent organs favoured a diagnosis of malignancy.

A comparison of ThinPrep Imager-assisted with manual screening, and its place in the New Zealand cervical cancer screening program.

AIMS: Overseas studies have shown equivalent sensitivity and specificity between computer-assisted screening and manual screening, with increased screener productivity. This study was undertaken to test these in a New Zealand laboratory setting. METHODS: A total of 9232 slides were read manually alone, and following ThinPrep Imager-assisted screening, and the results compared. Two senior screeners and a cytopathologist reviewed the slides with discordant results. RESULTS: The detection rate for abnormalities was 7.30% for Imager-assisted screening and 7.83% for manual screening. The concordance in diagnosis of abnormalities ranged from 72.7% to 100% with the lowest concordance for high-grade abnormalities diagnosed by Imager-assisted screening. The rate of unsatisfactory smears with Imager-assisted screening is half that of manual screening. There was a screener productivity increase of 140%. In all but one case, abnormal cells were identified by the Imager but screeners varied in their interpretations. CONCLUSIONS: Overall, Imager-assisted screening was as sensitive as manual screening, and more sensitive for high-grade lesions, with a halving of the rate of unsatisfactory smears. In the setting of the New Zealand cervical screening program, the initial screen by the Imager removes the need for a second, rapid rescreen required by the program for manual screening.

Salivary duct carcinoma arising in IgG4-related autoimmune disease of the parotid gland.

Hyper IgG4 disease or IgG4-related sclerosing/autoimmune disease is a multisystem condition characterized histologically by fibrosis, lymphoplasmacytic infiltration, and abundant IgG4 plasma cells associated with raised serum IgG4 levels. We present a case of salivary duct carcinoma of the parotid gland in a background of chronic sclerosing sialadenitis that also involved the submandibular gland with associated regional lymphadenopathy. The serology showed raised total IgG levels of 16.3 g/L (reference range, 6.0-15.0) and raised IgG4 levels of 3.41 g/L (reference range, 0.07-1.70). The salivary duct carcinoma contained areas of dense fibrosis and abundant IgG4-positive plasma cells (>100 per high-power field [hpf]). The adjacent noncarcinomatous areas, submandibular gland, and regional lymph nodes also contained plasma cells immunoreactive to IgG4 with densities higher than 100/hpf. To the best of our knowledge, this case is the first documentation of malignancy occurring in a background of IgG4-related autoimmune disease of the salivary gland.

Liver resection for hepatocellular carcinoma in a hepatitis B endemic area.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary malignancy of the liver. Treatment options include liver resection, tumor ablation, and liver transplantation. METHODS: We report the results of all patients undergoing partial hepatectomy for HCC with curative intent from a center where all major treatment modalities were available. RESULTS: A series of 53 patients were identified, of whom 72% had underlying liver disease, mostly chronic hepatitis B infection. Altogether, 57% of patients underwent major resections, of whom 43% had histologically proven cirrhosis. Postoperative morbidity and mortality occurred in 41.5% and 7.5%, respectively. After a median follow-up of 34 months, the survival probabilities at 1, 3, and 5 years were 74.1%, 54.1%, and 42.6%, respectively. A total of 47% developed recurrent disease over the study period with a median disease-free survival of 13.8 months. The probabilities of recurrence at 1, 3, and 5 years were 35.2%, 49.4%, and 55.9%, respectively. Among those who developed recurrence, 76% died, with a median time to death from the time the recurrence was diagnosed of 7.8 months. There was a good association between the CLIP score and survival following liver resection. Multivariate analysis showed that only tumor recurrence and the presence of cirrhosis was a significant determinant of the risk of tumor-related death. CONCLUSION: These findings confirm that with careful patient selection liver resection for HCC can achieve good long-term patient survival and acceptable risks.

Liver transplantation for hepatocellular carcinoma in New Zealand: a prospective intent-to-treat analysis.

UNLABELLED: Liver transplantation (LT) is potentially curative for early hepatocellular carcinoma (HCC) but time spent on the waiting list exposes patients to the risk of tumour progression prior to transplantation. AIMS: We prospectively evaluated the outcome for New Zealand patients listed for LT with a pre-transplant diagnosis of HCC. METHODS: Patients with 1 to 3 tumours, up to 5 cm in diameter, and no vascular invasion or extra-hepatic disease on imaging, were considered eligible for LT. The results were analysed by intention to treat from the time of listing. RESULTS: Fifty-nine patients were listed with a pre-transplant diagnosis of HCC between February 1998 and June 2004. Ten (17%) were de-listed before LT because of tumour progression, and 9 of 45 (20%) who underwent LT have experienced tumour recurrence up to 59 months post-transplant. For patients listed with a diagnosis of HCC, 5-year actuarial survival was 56.1% from the time of listing. For those listed and transplanted with a diagnosis of HCC, 5-year actuarial survival from the time of transplant was 63.5%. This is significantly worse than the 89.8% 5-year survival for patients transplanted without HCC over the same period (p=0.018) and this difference was entirely due to tumour recurrence. CONCLUSIONS: 37% of patients listed according to our criteria were either de-listed due to tumour progression or experienced recurrence after LT. Based on this experience strategies aimed at preventing waiting list drop out have been adopted, however expansion of tumour-related selection criteria cannot be recommended without a concomitant increase in donor organ availability.

Intragraft gene and protein expression in rat liver allografts treated with costimulatory blockade alone or in combination with CyA.

BACKGROUND: Costimulatory blockade has been shown to prevent acute rejection (AR) and promote long-term graft survival in a number of animal models including nonhuman primates. The effect of concomitant administration of conventional immunosuppressives on long-term liver allograft survival and intragraft expression of immune mediators has not previously been examined. MATERIALS AND METHODS: A high-responding Dark Agouti to Lewis orthotopic liver transplant (LEW OLT) model was used to compare anti-CD154 alone, or in combination with cyclosporin (CyA) on allograft survival. Donor-specific reactivity was assessed by mixed lymphocyte reaction (MLR) and allogeneic skin grafts. Surviving rats were euthanized on day 150 and intragraft gene (CD80, 86, 152, 154, IFN-gamma, IL-2, IL-6, IL-10, IL-13, TNF-alpha, TGF-beta, IL-7, Fas-ligand, Granzyme B, bax, and bcl(2)) and protein (CD4, CD8, ED1, CD154, CD80, CD86) expression was measured. RESULTS: Untreated control recipients had a median survival time of 5 days. Recipients treated with anti-CD154 survived to beyond 150 days with no evidence of AR. Concomitant administration of CyA did not alter the long-term survival. There was no difference in the serum aspartate aminotransferase between treatment groups or a change over time. All treated recipients showed a reduction in donor-specific MLR at day 40 and 60 but had persistence of donor reactivity to skin grafts at day 100. Histologically, liver architecture was well preserved despite the presence of a nondestructive mononuclear cell infiltrate. Analysis of intragraft gene expression revealed an inverse relationship between the duration of anti-CD154 therapy and the gene expression of costimulatory molecules and Th1 cytokine transcripts. The pro-apoptotic gene, bax, was increased in recipients treated with anti-CD154, but not CyA, compared with normal liver. CONCLUSIONS: These data demonstrate that anti-CD154 therapy either alone or in combination with CyA allows for the long-term survival of liver allografts in the rat despite there being a difference in the intragraft gene and protein profile.

Analysis of intragraft gene and protein expression of the costimulatory molecules, CD80, CD86 and CD154, in orthotopic liver transplant recipients.

CD40-CD154 and/or CD28-CD80/86 costimulatory blockade induces long-term allograft survival in numerous animal models. Studies examining the expression of costimulatory molecules during acute cellular rejection (ACR) have been limited to renal and cardiac allografts. The aim of this study was to describe the relationship between intragraft costimulatory molecule expression in OLT recipients and ACR. Forty-five liver biopsies were obtained at reperfusion and day 7. Gene and protein expression of CD80, CD86 and CD154 were analyzed by RT-PCR and immunohistochemistry. CD154 protein expression was present in 13 of 18 patients with a RAI score of 4, but in only two of 14 patients with a RAI score of <4. There was a strong association between the RAI score and the presence of CD80 and CD154 immunoreactivity. CD86 protein expression did not correlate with the severity of ACR. In reperfusion biopsies CD154, but not CD80 or CD86, protein expression correlated with the total ischaemic time. There was no association between expression of costimulatory molecule genes and ACR. In conclusion, we have demonstrated an association between CD154 and CD80 protein expression and ACR in orthotopic liver allografts.

Liver transplantation in New Zealand: the first four years.

AIM: To summarise the transplant-related activity of the New Zealand Liver Transplant Unit over the first four years. METHODS: The records of all patients assessed for liver transplantation between 1 December 1997 and 30 December 2001 were examined. Listing criteria, demographics, waiting time, transplant-hospitalisation details and long-term outcome for those who underwent liver transplantation were recorded. RESULTS: One hundred and eighty six patients over 14 years of age (acute liver failure 28, chronic liver disease 158) were assessed and 150 were listed for liver transplantation. Fifteen died waiting, 13 were de-listed (6 for cancer progression) and 14 remain listed. One hundred and nine liver transplants (including 1 combined heart-liver, 1 sequential liver-bone marrow and 5 re-transplants) were performed on 104 patients (13 acute liver failure, 96 chronic liver failure or hepatocellular carcinoma). The median waiting time was 2 days (range 0 5) for acute liver failure and 62 days (range 0 320) for other patients. Median age at transplant was 50 years (range 14-70); 73 patients (66%) were male; 71 (65%) were European; 13 (12%) Maori; 12 (11%) Pacific Islander; and 8 (7%) Asian. Median duration of surgery was 480 minutes (range 300 720 minutes); red cell transfusion 5 units (0 32); intensive care and total hospital stays were 2 (range 1 17) and 11 days (range 6 91). One transplanted patient died in hospital, 1- and 3-year patient survival was 94% and 87% and corresponding graft survival was 91% and 83%. Ninety three transplanted patients (89%) are alive. Of the 92 patients at least three months post-transplant, 62 (67%) were employed. CONCLUSION: Liver transplantation is now established in New Zealand as treatment of choice for acute and chronic liver failure and small hepatocellular carcinoma. Excellent outcomes have been attained in those transplanted to date. Reduction in waiting list mortality will require identification of and investment in strategies that will expand the donor organ availability.

Costimulatory blockade prevents early rejection, promotes lymphocyte apoptosis, and inhibits the upregulation of intragraft interleukin-6 in an orthotopic liver transplant model in the rat.

Costimulatory pathways have a pivotal role in the T-cell response to alloantigen. The role of costimulatory blockade with anti-CD154 in orthotopic liver transplantation (OLT) has not been examined previously. This study aims to investigate effects of anti-CD154 and CTLA4-immunoglobulin (Ig) in the early post-OLT period using a major histocompatibility complex-disparate fully arterialized OLT model in the rat. Lewis rats underwent OLT with Dark Agouti liver allografts. Recipients were randomized to receive (1) isotype control, (2) anti-CD154, (3) CTLA4-Ig, or (4) cyclosporine A (CyA). Rats were killed day 8, and specimens were obtained for histological examination, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling, immunohistochemistry, and quantitative reverse-transcriptase polymerase chain reaction. An additional five transplant recipients were treated with anti-CD154 for 14 days postoperatively to assess long-term allograft survival. All isotype control animals died on or before day 6 of acute rejection. Apart from four deaths caused by nonimmunologic causes, all treated recipients survived to day 8. The median survival of rats treated for 14 days with anti-CD154 was greater than 150 days. Serum aspartate aminotransferase and bilirubin levels normalized by day 3 in the CyA group and day 5 in transplant recipients treated with costimulatory blockade. Histologically, there was no difference between isotype controls and CTLA4-Ig-treated animals, whereas anti-CD154-treated transplant recipients had a lower Banff score. CD4+ and CD8+ T-cell infiltrates were prominent in transplant recipients treated with costimulatory blockade. Intragraft analysis showed an increase in lymphocyte apoptosis, Fas ligand messenger RNA expression, and reduction in interleukin-6 gene expression in transplant recipients treated with costimulatory blockade. Costimulatory blockade did not alter intragraft gene expression of other mediators of T-cell priming, differentiation, and effector function compared with isotype control animals. In conclusion, costimulatory blockade prevented acute rejection, enabled long-term survival, and increased intragraft lymphocyte apoptosis in a high-responding rat OLT model.