When and how do patients with cardiac amyloidosis die?

BACKGROUND: Cardiac amyloidosis (CA) is an underappreciated cause of morbidity and mortality. Light-chain (AL) and transthyretin (ATTR) amyloidosis have different disease trajectories. No data are available on subtype-specific modes of death (MOD) in patients with CA. METHODS AND RESULTS: We retrospectively investigated 66 with AL and 48 with wild-type ATTR amyloidosis (ATTRwt) from 2000 to 2018. ATTRwt differed from AL by age (74.6 ± 5.4 years vs. 63 ± 10.8 years), posterior wall thickness (16.8 ± 3.3 mm vs. 14.3 ± 2.2 mm), left ventricular mass index (180.7 ± 63.2 g/m2 vs. 133.5 ± 42.2 g/m2), and the proportions of male gender (91.7% vs. 59.1%), atrial enlargement (92% vs. 68.2%) and atrial fibrillation (50% vs. 12.1%). In AL NYHA Functional Class and proteinuria (72.7% vs. 39.6%) were greater; mean arterial pressure (84.4 ± 13.5 mmHg vs. 90.0 ± 11.3 mmHg) was lower. Unadjusted 5-year mortality rate was 65% in AL-CA vs. 44% in the ATTRwt group. Individuals with AL-CA were 2.28 times ([95%CI 1.27-4.10]; p = 0.006) more likely to die than were individuals with ATTRwt-CA. Information on MOD was available in 56 (94.9%) of 59 deceased patients. MOD was cardiovascular in 40 (66.8%) and non-cardiovascular in 16 (27.1%) patients. Cardiovascular [28 (68.3%) vs. 13 (80%)] death events were distributed equally between AL and ATTRwt (p = 0.51). CONCLUSION: Our data indicate no differences in MOD between patients with AL and ATTRwt cardiac amyloidosis despite significant differences in clinical presentation and disease progression. Cardiovascular events account for more than two-thirds of fatal casualties in both groups.

Klotho is upregulated in human cardiomyopathy independently of circulating Klotho levels.

Klotho is an antiaging protein which exerts known cardioprotection. In kidney, trans-membrane Klotho acts as essential co-receptor of fibroblast growth factor 23 (FGF23). In the heart, soluble Klotho (sKlotho) protects from systolic dysfunction independently of FGF23. Here, we analyzed the association of FGF23 and sKlotho upon progression of chronic heart failure (CHF) and analyzed Klotho expression in human hearts. Serum levels of sKlotho and FGF23 were measured in 287 patients with cardiomyopathy (CMP). Tissue samples from CMP (n = 10) and healthy control hearts (n = 10) were analyzed for Klotho mRNA and protein expression. Individuals in the first FGF23 tertile were 4.1 times more likely of freedom from death, heart transplantation or assist device implantation compared to third tertile. No relationship was found between sKlotho and the combined endpoint. Instead, Klotho mRNA encoding the full-length form was upregulated in human CMP hearts. Immunoblotting confirmed upregulation of sKlotho associated with increased expression of proteases involved in cleavage of Klotho suggesting rather local effects of Klotho in the heart. Therefore, we conclude that in contrast to FGF23, serum sKlotho is not associated with disease severity or progression in CHF. Instead, Klotho is expressed and upregulated in diseased hearts, suggesting local paracrine effects.

COX-2 expression in highly aggressive thyroid malignancies - indication for a possible therapeutic option?

Both anaplastic thyroid carcinoma (ATC) and angiosarcoma of the thyroid (AST) are highly aggressive malignancies with very limited therapeutic options. Since selective inhibition of COX-2, for example, by celecoxib has been shown to suppress both tumour formation and progression, we investigated COX-2 protein expression in a series of ATC and AST (26 cases each) using immunohistochemistry. COX-2 expression was demonstrated in 13 ATC (50%) and 11 AST (42%); a strong COX-2 expression in more than 50% of vital tumour cells was found in 5 ATC and 5 AST, respectively. Although a recently performed phase II trial applying celecoxib failed overall to halt tumour progression in differentiated thyroid carcinoma, the two cases with partial or complete remission noted in this study were related to tumours with immunohistochemically proven strong COX-2 expression. The strong COX-2 expression observed in approximately 20% of our ATC and AST samples may thus indicate selective patients with a possible therapeutic option for an otherwise fatal disease.

EpCAM is predominantly expressed in high grade and advanced stage urothelial carcinoma of the bladder.

BACKGROUND: EpCAM is an adhesion molecule of the basolateral membranes in a variety of epithelial cells. Over-expression has been detected in many epithelial tumours and has been associated with high stage, high grade and a worse survival in some tumour types. AIMS: To assess the prognostic value of EpCAM in urothelial carcinoma of the bladder. METHODS: EpCAM expression was analysed by immunohistochemistry using a monoclonal antibody (clone VU-1D9) on a tissue microarray comprising 99 urothelial carcinomas of the bladder diagnosed between 1994 and 1997. RESULTS: A significant relationship between high grade, advanced stage, and EpCAM expression was found, and expression of EpCAM was associated with a worse overall survival when compared to EpCAM negative tumours (p = 0.033). Multivariate analysis showed that EpCAM expression was not an independent prognostic factor for overall survival in urothelial carcinoma of the bladder. CONCLUSION: EpCAM expression is associated with advanced stage, high grade and poor overall survival in urothelial carcinoma of the bladder, but lacks an independent prognostic significance. The strong association with high grade tumours suggests a possible role during tumour progression and makes EpCAM a potential target for antibody mediated therapy.

Expression and prognostic significance of Tetranectin in invasive and non-invasive bladder cancer.

Tetranectin (TN) is a plasminogen kringle-4 binding protein that can be detected in the plasma and the extracellular matrix. In malignancies, TN is thought to enhance proteolytic processes enabling tumor cells to invade and metastasize. So far, TN expression has not been described in transitional cell bladder cancer (TCC), and there is no information on the prognostic significance of its in situ expression. TN expression was studied in 99 TCC patients diagnosed between 1994 and 1997. Immunohistochemistry was performed on a tissue microarray using a monoclonal antibody against TN (clone 11F1). Within the mean follow-up period of 60 months, pTa and pT2-4 TCC patients with stainable TN in the tumor stroma had a significant shorter recurrence-free survival and higher risk of recurrence compared to those without stainable TN (p = 0.0002 for both). TN expression in the tumor cells did not influence recurrence-free survival. In conclusion, TN is expressed in a subgroup of bladder cancer patients with a higher risk of recurrence who may take benefit from a closer follow-up.

Evidence for Somatostatin receptor 2 in thyroid tissue.

Somatostatin receptor scintigraphy has found considerable interest for imaging thyroid tumours. Recently, also therapeutic application of Somatostatin analogues labelled with beta-emitting radionuclides has been suggested as treatment option for thyroid tumours with absent radioiodine uptake. Most of the radiolabelled analogues available show a predominant affinity for Somatostatin receptor subtype 2. This study reports on the in vitro characterisation of Somatostatin receptor subtype mRNAs in thyroid tumours and normal thyroid tissue by means of RT-PCR. Surgical samples of 21 patients were collected, and mRNA of 16 tumour and 17 control specimen was isolated. mRNA expression for Somatostatin, SSTR subtype 1-5, thyroid markers (NIS, TSH, Tg, TPO) and control markers (GAPDH, beta-actin) was determined. PCR results were correlated with immunohistochemistry staining using SSTR2 receptor specific antibodies. 94% of all samples expressed Somatostatin receptor mRNA with predominant expression of subtype 2, less predominant of subtype 5 and subtype 3. Somatostatin receptor subtype 2 mRNA expression correlated well with immunohistochemical staining pattern in 13/16 samples, SSTR2 immunohistochemistry was positive in 87% of the samples. Our results show that Somatostatin receptor 2 is predominantly expressed on thyroid tissue and is a valid target for treatment of thyroid tumours. Octreotide derivatives currently used in Nuclear medicine seem to be well suited to target receptors expressed in thyroid tumours.

The C allele of the GNB3 C825T polymorphism of the G protein beta3-subunit is associated with an increased risk for the development of oncocytic thyroid tumours.

Carriers of the C allele of the common C825T polymorphism in the GNB3 gene of the G protein have been associated with the development of follicular thyroid adenomas. Since the C allele of this polymorphism is related to a lower signalling capacity, it was speculated whether the C825T polymorphism may play a particular role in oncocytic thyroid tumours, which are recognized for their reduced ability to synthesize thyroid-specific proteins and hormones, although they possess an intact thyroid-stimulating hormone receptor-adenylyl cyclase system. Using pyrosequencing, both the genotype distribution and the allele frequency of the C825T polymorphism were investigated in a series of 104 patients with oncocytic thyroid tumours of follicular cell origin [58 adenomas, 41 follicular thyroid carcinomas (FTCs), and five papillary thyroid carcinomas (PTCs)]; the results were compared with those obtained from 321 age and gender-matched healthy blood donors and a series of 327 non-oncocytic thyroid tumours of follicular cell origin (119 adenomas, 80 FTCs, and 186 PTCs). Analysis of the genotype distribution (comparing oncocytic with non-oncocytic tumours of the present series) revealed a significantly increased odds ratio (OR) for CC versus TT (OR = 4.22; p = 0.011) and CC versus CT (OR = 1.62; p = 0.049) carriers to develop an oncocytic thyroid tumour; ORs to develop an oncocytic thyroid tumour were also increased comparing the genotype distribution between the group of oncocytic tumours and healthy controls for CC versus TT (OR = 3.73; p = 0.017) and CC versus all T carriers (OR = 1.56; p = 0.034). Oncocytic thyroid tumours as a group showed a statistically significant increase of the C-allele frequency when compared with all non-oncocytic tumours (p = 0.0039) as well as healthy blood donors (p = 0.017). The results strongly suggest that the C allele of the GNB3 C825T polymorphism of the G protein beta3-subunit is associated with an increased risk for the development of oncocytic thyroid tumours. This polymorphism may thus be considered a (co)factor favouring the development of oncocytic thyroid tumours, although the biological mechanism(s) underlying this association remain obscure.

Tetranectin in cerebrospinal fluid of patients with multiple sclerosis.

OBJECTIVE: Tetranectin (TN) is a glycoprotein and C-type lectin thought to play a prominent role in tissue remodelling. The aim of this study was to determine the TN serum and cerebrospinal fluid (CSF) concentration in patients with multiple sclerosis (MS) and controls. MATERIAL AND METHODS: Two-hundred-and-four patients, divided into four diagnostic groups, i.e. definite MS (n = 76), possible onset symptoms of MS (n = 48), other non-inflammatory neurological diseases (n = 61) and other inflammatory neurological diseases (n = 19) and 47 controls with no history of neurological disease were analysed for TN in serum and CSF using a polyclonal sandwich ELISA. RESULTS: All tested groups, e.g. definite MS, possible onset symptoms of MS, other neurological disease, both inflammatory and non-inflammatory, had decreased concentrations of TN in the CSF compared to the concentrations in controls. The quotient of TN in CSF divided by the concentration in serum (QTN) correlated significantly with the same quotient of albumin (QALB), was significantly correlated with the same quotient of albumin QALB. To account for differences in blood brain barrier permeability, we calculated a TN-index defined as: TN-index = QTN/QALB. QTN was significantly decreased in all groups compared to that in controls. However, in definite MS and patients with first attack of MS, the TN-index was not significantly different from that of controls. In contrast, other neurological diseases, both inflammatory and non-inflammatory, were associated with a decreased TN-index. CONCLUSION: These results indicate that TN may play a role in neurological diseases and may serve as a diagnostic aid in MS.

Aberrant tetranectin expression in human breast carcinomas as a predictor of survival.

AIMS: Tetranectin (TN), a plasminogen kringle 4 binding protein, is thought to play a prominent role in the regulation of proteolytic processes via binding to plasminogen. The aim of this study was to evaluate the expression of TN in human breast cancer and adjacent normal breast tissue and to determine the impact of this expression on survival. METHODS: A retrospective analysis was performed on 189 patients with breast cancer, with a median follow up time of 10.6 years. The expression of TN was assessed in tumour tissue and adjacent normal breast tissue by immunohistochemistry, and the prognostic relevance of its expression in tumour cells was evaluated. RESULTS: TN was highly expressed in connective tissue fibres surrounding normal breast epithelium, but not in normal epithelial cells. High expression of TN in tumour cells was found in 131 (69%) of the tumour samples. By western blot analysis, no significant difference in the amount and molecular weight of TN was seen between tumour tissue and normal tissue. Strong TN immunoreactivity in tumour tissue was predictive of poor disease free and tumour specific overall survival. By multivariate analysis, high TN expression in cancer cells was an independent prognostic factor for disease free and tumour specific overall survival. CONCLUSIONS: Our results demonstrate differential TN expression in normal and malignant breast tissue and a prognostic impact of TN protein expression in breast carcinoma tissue. These data suggest a possible role of TN in invasiveness and the metastatic spread of human breast cancer.

Epidermal growth factor receptor as a novel therapeutic target in anaplastic thyroid carcinomas.

Anaplastic thyroid carcinoma (ATC) is one of the most aggressive human malignancies, with a median survival of up to 6 months. Such a bad prognosis under the present treatment procedures suggests the need for novel approaches in the management of this disease. Since some epidermal growth factor receptor (EGFR) inhibitors are now in clinical trials and few data are available concerning EGFR expression in anaplastic thyroid carcinomas, we tried to estimate a possible overexpression of this receptor in a larger tumor series. Twenty-five ATCs, including 3 ATCs with poorly differentiated thyroid carcinoma (PDTC) parts, were immunohistochemically investigated with a mouse monoclonal antibody directed against EGFR (EGFR pharmDX kit). The tumors revealed primarily a distinct membranous staining pattern, and in several tumor cells an additional cytoplasmic reactivity could be observed. The anaplastic carcinomas presented with 5 of 25 (20%) without EGFR reaction, 10 of 25 (40%) with reactivity, and 10 of 25 (40%) with overexpression of the receptor. All ATCs with PDTC parts (100%) showed EGFR overexpression. Cytoplasmic reactivity was observed in 56% of all ATCs. A significant correlation was calculated for EGFR overexpression and cytoplasmic staining (P = 0.036). Concerning receptor overexpression, ATCs were significantly different from ATCs with PDTC parts (P = 0.023). For the first time, we present EGFR overexpression in ATC in a larger tumor series, demonstrating that EGFR overexpression is a common finding in ATC. For at least one-third of all anaplastic thyroid carcinomas, EGFR seems to be a promising agent for the targeted molecular therapy of these extraordinarily aggressive tumors.

[Thyroid tumors]. / Tumoren der Schilddrüse.

The vast majority of thyroid tumours are epithelial. In contrast to thyroid adenoma, which is a common tumour, thyroid carcinomas make up only 1% of all human malignancies. Routine pathology is regularly confronted with a differential diagnosis involving thyroid adenoma and carcinoma, as well as particular variants of these tumours. This paper deals with the standardised gross and histological examination, as well as the impact of immunohistochemistry and intraoperative frozen sections on thyroid pathology.

Her2/neu overexpression in differentiated thyroid carcinomas predicts metastatic disease.

To investigate the prognostic value of Her2/neu expression in differentiated thyroid carcinomas 103 patients were retrospectively investigated. All of them received surgical and an identical follow-up treatment. The patients with papillary and follicular thyroid cancer were further separated into two groups concerning their clinical development, including one group without distant metastasis (follow-up of minimum 8 years). The second group presented with distant metastases as a sign of an aggressive behaviour. Her2/neu was immunohistochemically detected on sections from formalin-fixed, paraffin-embedded tissues using c-erbB-2/Her-2/neu oncoprotein Ab-17 monoclonal antibody (mAb). In statistical analysis using the Mann-Whitney U-test and chi(2) test, Her2/neu protein overexpression was significantly correlated with prognosis. Both tumour entities without distant metastases showed significantly less cytoplasmic immunostaining than patients with development of metastases. Concerning the clinical outcome, Her2/neu overexpression may be regarded as a prognostic factor in differentiated thyroid carcinomas. Moreover, in addition to standard radio-iodine elimination therapy, application of Herceptin could lead to new successful therapeutic concepts for a number of patients with progressive thyroid cancer.

[Pathological examination of breast biopsy specimens]. / Pathologische Aufarbeitung von Mammabiopsaten.

No differences for long-term disease free survival could be found between breast conserving surgery and mastectomy. Most importantly is the fact that this therapy presents a significantly higher risk for local recurrence. The characterisation of this risk is one of the most important things to do. These findings result in a widespread change in treatment of breast cancer patients. Consequently an increase in interdisciplinary working between radiologists, surgeons and pathologists could be found. Histological examinations are necessary for diagnosis and exactly evaluation of the tumor extension. Microscopic evaluation of the resection margin is of most important interest, because there is a direct connection between local recurrence and tumor infiltration of the resection margin. We performed our investigations by the use of a standardized complete embedding method with the possibility of three-dimensional reconstruction, on a cohort of 280 patients. Additionally this method allowed the detection of all relevant findings on one hand and on the other hand an evaluation of all resection margins. Our results showed a breast conserving therapy including tumor free margins was performed in 67% of the patients. But there was a second resection necessary in 57% of the cases. An extensive tumor distribution as the detection of multifocal tumor spread was the reason for mastectomy in 33%. Our findings point out the necessarily of the histological examination in the line of the complete embedding method of the breast biopsy material in order to analyse the tumor including resection margin evaluation.

Elevated plasma levels of crosslinked fibrinogen gamma-chain dimer indicate cancer-related fibrin deposition and fibrinolysis.

Cancer-related fibrin deposition and fibrinolysis were investigated by two-dimensional gel electrophoresis of human solid tumor and effusion specimen in addition to plasma samples. Fibrinogen gamma-chain dimer indicating fibrin deposition and plasmin-generated fibrinogen beta-chain fragments were identified in various solid tumor types by amino acid sequencing, mass spectrometry analysis and Western blotting. In tumor-associated effusions, these techniques allowed to observe plasmin-generated fragments of fibrinogen alpha, beta and gamma-chains in addition to elevated levels of acute-phase proteins. Similar observations were made in case of inflammation-associated effusions. No fibrin degradation product was observed in plasma samples, however, high amounts of fibrinogen gamma-chain dimer crosslinked by transglutaminase were detected in plasma from tumor patients, but not in plasma from controls and patients suffering acute infections and/or inflammations. This finding demonstrated that high transglutaminase activity may be associated with cancer. The presented data indicate that the amount of crosslinked fibrinogen gamma-chain dimer in plasma may correlate with tumor-associated fibrin deposition. The tumor-biological relevance of this potential marker protein is discussed.

Quantification and characterization of pregnancy-associated complexes of angiotensinogen and the proform of eosinophil major basic protein in serum and amniotic fluid.

BACKGROUND: The proform of eosinophil major basic protein (ProMBP) exists in serum from pregnant women complexed with a variable fraction of angiotensinogen (Ang). A subfraction further binds complement C3dg in a 2:2:2 complex. The function, physiology, and clinical importance of ProMBP complexes are unknown, and the specific quantification of these complexes has not been possible. METHODS: We developed an ELISA for the ProMBP/Ang complexes, using a monoclonal antibody against ProMBP for capture and a chicken anti-human Ang antiserum for detection. Calibrators were standardized with WHO IRP 78/610 for pregnancy proteins in the assay range 0.95-15.6 mIU/L. RESULTS: The concentrations of ProMBP/Ang complexes in serum of nonpregnant blood donors (n = 79) were log-normally distributed with a central 95th interval of 985-3655 mIU/L. In pregnancy, mean serum concentrations were increased from week 7, and the concentrations reached term concentrations in week 18. ProMBP/Ang complexes eluted in gel filtration as a broad peak with a molecular mass of approximately 230 kDa. The concentration of ProMBP/Ang/C3dg increased during blood coagulation, suggesting that the ProMBP/Ang/C3dg complex may be a marker of complement activation. CONCLUSIONS: ProMBP/Ang complexes are present in serum from nonpregnant persons as well as pregnant women, and the direct assays described here will make it possible to study the biochemistry and the clinical significance of different ProMBP complexes in pathological conditions and pregnancy.

A human common nuclear matrix protein homologous to eukaryotic translation initiation factor 4A.

Amino acid sequencing and mass spectrometry revealed identity of a human nuclear matrix protein, termed hNMP 265, with a predicted protein of gene KIAA0111. Two-dimensional electrophoresis and Northern hybridization showed the protein to ubiquitously occur in various human cell types. Exhibiting DEAD-box motifs characteristic for RNA helicases, hNMP 265 is highly similar to the human initiation factors eIF4A-I and -II. On the other hand, hNMP 265 greatly differs from the initiation factors by a N-terminal sequence rich in charged amino acids. Sequence searches and alignments indicate proteins related to hNMP 265 in other eukaryotes. Chimeras between hNMP 265 and green fluorescence protein or hapten appeared as speckles in extranucleolar regions in the nucleus, but not in the cytoplasm. Experiments with tagged deletion mutants indicated that the N-terminal amino acid sequence is necessary for nuclear localization. A putative role of hNMP 265 in pre-mRNA processing is discussed.

"Atypical" medullary thyroid carcinoma with little or no calcitonin expression.

In a retrospective analysis of 142 medullary thyroid carcinomas, four sporadic cases with an unusual histological and immunohistochemical appearance were found. Three cases (two males, one female) had very few calcitonin-positive tumour cells, while the fourth case (male) completely lacked calcitonin immunoreactivity at both mRNA and protein levels, whereas a variety of neuroendocrine markers were positive in at least 50% of tumour cells. The four tumours were completely devoid of carcinoembryonic antigen expression and of amyloid. Differential diagnosis and histogenetic considerations are discussed.

Sonographic appearance of an appendix carcinoma.

Malignant appendix tumours are rare entities. Especially adenocarcinomas, which only appear in about 10% of appendix tumours, are very seldom. Preoperative diagnosis is very difficult due to a lack of typical clinical signs and a clinical appearance mimicking perforated appendicitis. Nevertheless, sonography is able to show indirect signs and therefore it can provide the surgeon with more information for a better operative treatment.