Biopsy of the facial nerve in slow-onset facial palsy.

INTRODUCTION: Primary squamous cell carcinoma of the parotid gland typically presents as a palpable, often painless mass. Peripheral facial palsy as the only sign of malignant neoplasia is rare. In these cases, the diagnosis is regularly confirmed by radiological imaging followed by surgical exploration and biopsy. However, if there is no detection of malignant lesions and no evidence of a tumor, the reluctance to take a biopsy of an unremarkable nerve can lead to misdiagnoses. CASE REPORT: A 40-year-old female patient without medical history presented to our clinic with a complete right-sided peripheral facial palsy that had slowly progressed for 2.5 years. All other otorhinolaryngological examination findings were within normal limits. Magnetic resonance imaging examination of the head and neck and 18-fluorodeoxyglucose positron emission tomography showed unremarkable results. We proceeded with surgical exploration, which revealed no evidence of a tumor and an externally completely unremarkable facial nerve. A biopsy from the main trunk area of the nerve revealed an infiltration by a squamous cell carcinoma. Total parotidectomy with resection and reconstruction of the facial nerve and neck dissection was performed. Considering the absence of a primary tumor and other tumor formations the diagnosis of a completely regressive primary squamous cell carcinoma of the parotid gland was confirmed. CONCLUSION: In conclusion, in the case of slow-onset peripheral facial palsy that persists without signs of recovery, a gadolinium-enhanced MRI should be performed. If imaging is unremarkable and there is no primary tumor detection along the course of the facial nerve, a surgical exploration with biopsy of the facial nerve is necessary.

Adult intracranial ependymoma-relevance of DNA methylation profiling for diagnosis, prognosis, and treatment.

BACKGROUND: A methylation-based classification of ependymoma has recently found broad application. However, the diagnostic advantage and implications for treatment decisions remain unclear. Here, we retrospectively evaluate the impact of surgery and radiotherapy on outcome after molecular reclassification of adult intracranial ependymomas. METHODS: Tumors diagnosed as intracranial ependymomas from 170 adult patients collected from 8 diagnostic institutions were subjected to DNA methylation profiling. Molecular classes, patient characteristics, and treatment were correlated with progression-free survival (PFS). RESULTS: The classifier indicated an ependymal tumor in 73.5%, a different tumor entity in 10.6%, and non-classifiable tumors in 15.9% of cases, respectively. The most prevalent molecular classes were posterior fossa ependymoma group B (EPN-PFB, 32.9%), posterior fossa subependymoma (PF-SE, 25.9%), and supratentorial ZFTA fusion-positive ependymoma (EPN-ZFTA, 11.2%). With a median follow-up of 60.0 months, the 5- and 10-year-PFS rates were 64.5% and 41.8% for EPN-PFB, 67.4% and 45.2% for PF-SE, and 60.3% and 60.3% for EPN-ZFTA. In EPN-PFB, but not in other molecular classes, gross total resection (GTR) (P = .009) and postoperative radiotherapy (P = .007) were significantly associated with improved PFS in multivariable analysis. Histological tumor grading (WHO 2 vs. 3) was not a predictor of the prognosis within molecularly defined ependymoma classes. CONCLUSIONS: DNA methylation profiling improves diagnostic accuracy and risk stratification in adult intracranial ependymoma. The molecular class of PF-SE is unexpectedly prevalent among adult tumors with ependymoma histology and relapsed as frequently as EPN-PFB, despite the supposed benign nature. GTR and radiotherapy may represent key factors in determining the outcome of EPN-PFB patients.

Functional Implications of LH/hCG Receptors in Pregnancy-Induced Cushing Syndrome.

CONTEXT: Elevated human choriogonadotropin (hCG) may stimulate aberrantly expressed luteinizing hormone (LH)/hCG receptor (LHCGR) in adrenal glands, resulting in pregnancy-induced bilateral macronodular adrenal hyperplasia and transient Cushing syndrome (CS). OBJECTIVE: To determine the role of LHCGR in transient, pregnancy-induced CS. DESIGN SETTING PATIENT AND INTERVENTION: We investigated the functional implications of LHCGRs in a patient presenting, at a tertiary referral center, with repeated pregnancy-induced CS with bilateral adrenal hyperplasia, resolving after parturition. MAIN OUTCOME MEASURES AND RESULTS: Acute testing for aberrant hormone receptors was negative except for arginine vasopressin (AVP)-increased cortisol secretion. Long-term hCG stimulation induced hypercortisolism, which was unsuppressed by dexamethasone. Postadrenalectomy histopathology demonstrated steroidogenically active adrenocortical hyperplasia and ectopic cortical cell clusters in the medulla. Quantitative polymerase chain reaction showed upregulated expression of LHCGR, transcription factors GATA4, ZFPM2, and proopiomelanocortin (POMC), AVP receptors (AVPRs) AVPR1A and AVPR2, and downregulated melanocortin 2 receptor (MC2R) vs control adrenals. LHCGR was localized in subcapsular, zona glomerulosa, and hyperplastic cells. Single adrenocorticotropic hormone-positive medullary cells were demonstrated in the zona reticularis. The role of adrenal adrenocorticotropic hormone was considered negligible due to downregulated MC2R. Coexpression of CYP11B1/CYP11B2 and AVPR1A/AVPR2 was observed in ectopic cortical cells in the medulla. hCG stimulation of the patient's adrenal cell cultures significantly increased cyclic adenosine monophosphate, corticosterone, 11-deoxycortisol, cortisol, and androstenedione production. CTNNB1, PRKAR1A, ARMC5, and PRKACA gene mutational analyses were negative. CONCLUSION: Nongenetic, transient, somatic mutation-independent, pregnancy-induced CS was due to hCG-stimulated transformation of LHCGR-positive undifferentiated subcapsular cells (presumably adrenocortical progenitors) into LHCGR-positive hyperplastic cortical cells. These cells respond to hCG stimulation with cortisol secretion. Without the ligand, they persist with aberrant LHCGR expression and the ability to respond to the same stimulus.

Glutamate enrichment as new diagnostic opportunity in breast cancer.

Exogenous glutamine is an important source of energy and molecular building blocks for many tumors. There is a renewed interest in therapeutically targeting glutamine metabolism due to the recent discovery of two novel glutaminase inhibitors. To quantify the dysregulation of the glutamate-glutamine equilibrium in breast cancer, metabolomics analysis of 270 clinical breast cancer samples and 97 normal breast samples was carried out using gas chromatography combined with time-of-flight mass spectrometry. Positive correlation between glutamate and glutamine in normal breast tissues switched to negative correlation between glutamate and glutamine in breast cancer tissues. Compared with the ratio of glutamate to glutamine in normal tissues, we found 56% of the ER+ tumor tissues and 88% of the ER- tumor tissues glutamate-enriched. The glutamate-to-glutamine ratio (GGR) significantly correlated with ER status (p = 8.0E-09) and with tumor grade (p = 3.3E-05). Higher levels of GGR were associated with prolonged overall survival in univariate analysis (HR = 0.77, p = 0.027) and in multivariate analysis (HR = 0.73, p = 0.038). GGR levels were reflected in an unsupervised clustering of metabolomics profiles. In a supervised analysis of metabolomics data and of genome-wide expression data, replacement of GGR by metabolite surrogate markers was feasible, while replacement of GGR by RNA markers had a limited accuracy. Functional analysis of the gene expression data showed negative correlation between glutamate enrichment and activation of peroxisome proliferator-activated receptor (PPAR) pathway. Our findings may have important implications for patient stratification related to utilization of glutaminase inhibitors.

Higher cytoplasmic and nuclear poly(ADP-ribose) polymerase expression in familial than in sporadic breast cancer.

Poly(ADP-ribose) polymerase 1 (PARP) is a key element of the single-base excision pathway for repair of DNA single-strand breaks. To compare the cytoplasmic and nuclear poly(ADP-ribose) expression between familial (BRCA1, BRCA2, or non BRCA1/2) and sporadic breast cancer, we investigated 39 sporadic and 39 familial breast cancer cases. The two groups were matched for hormone receptor status and human epidermal growth factor receptor 2 status. Additionally, they were matched by grading with a maximum difference of ±1 degree (e.g., G2 instead of G3). Cytoplasmic PARP (cPARP) expression was significantly higher in familial compared to sporadic breast cancer (P = 0.008, chi-squared test for trends) and a high nuclear PARP expression (nPARP) was significantly more frequently observed in familial breast cancer (64 %) compared with sporadic breast cancer (36 %) (P = 0.005, chi-squared test). The overall PARP expression was significantly higher in familial breast cancer (P = 0.042, chi-squared test). In familial breast cancer, a combination of high cPARP and high nPARP expression is the most common (33 %), whereas in sporadic breast cancer, a combination of low cPARP and intermediate nPARP expression is the most common (39 %). Our results show that the overall PARP expression in familial breast cancer is higher than in sporadic breast cancer which might suggest they might respond better to treatment with PARP inhibitors.

Novel theranostic opportunities offered by characterization of altered membrane lipid metabolism in breast cancer progression.

Activation of lipid metabolism is an early event in carcinogenesis and a central hallmark of many cancers. However, the precise molecular composition of lipids in tumors remains generally poorly characterized. The aim of the present study was to analyze the global lipid profiles of breast cancer, integrate the results to protein expression, and validate the findings by functional experiments. Comprehensive lipidomics was conducted in 267 human breast tissues using ultraperformance liquid chromatography/ mass spectrometry. The products of de novo fatty acid synthesis incorporated into membrane phospholipids, such as palmitate-containing phosphatidylcholines, were increased in tumors as compared with normal breast tissues. These lipids were associated with cancer progression and patient survival, as their concentration was highest in estrogen receptor-negative and grade 3 tumors. In silico transcriptomics database was utilized in investigating the expression of lipid metabolism related genes in breast cancer, and on the basis of these results, the expression of specific proteins was studied by immunohistochemistry. Immunohistochemical analyses showed that several genes regulating lipid metabolism were highly expressed in clinical breast cancer samples and supported also the lipidomics results. Gene silencing experiments with seven genes [ACACA (acetyl-CoA carboxylase α), ELOVL1 (elongation of very long chain fatty acid-like 1), FASN (fatty acid synthase), INSIG1 (insulin-induced gene 1), SCAP (sterol regulatory element-binding protein cleavage-activating protein), SCD (stearoyl-CoA desaturase), and THRSP (thyroid hormone-responsive protein)] indicated that silencing of multiple lipid metabolism-regulating genes reduced the lipidomic profiles and viability of the breast cancer cells. Taken together, our results imply that phospholipids may have diagnostic potential as well as that modulation of their metabolism may provide therapeutic opportunities in breast cancer treatment.

Fibrosis progression after liver transplantation in patients with recurrent hepatitis C.

BACKGROUND/AIMS: Aim of our study was to analyze fibrosis progression after liver transplantation (OLT) in hepatitis C virus (HCV)-infected patients based on protocol liver biopsies and to identify risk factors, which may play a role in the development of severe fibrosis stages. METHODS: One hundred and eighty-three liver graft recipients who had a histological follow-up evaluation of 1 year after OLT were analyzed. Overall 1039 protocol liver biopsies were performed after 1-, 3-, 5-, 7- and 10 years and staged according to the Scheuer score. RESULTS: The fibrosis progression rate was not linear. The fibrosis scores were 1.2 after one, 1.7 after three, 1.9 after five, 2.1 after 7 and 2.2 after 10 years. The 39 recipients with fibrosis stages 3 or 4 in the 1-year biopsy had a significantly reduced survival rate, while fibrosis stage 0-2 indicated excellent survival. Independent risk factors for progression of fibrosis at 1 year were HCV genotype 1 and 4 (P=0.01) and donor age>33 years (P=0.01), whereas risk factors for development of cirrhosis (30/183 recipients (16%)) were donor age (P=0.002) and multiple steroid pulses (P=0.05). CONCLUSIONS: These data provide information on the course of recurrent hepatitis C and may be helpful to individualize the treatment of transplanted patients.

Hepatocellular adenoma and focal nodular hyperplasia after long-term use of danazol for endometriosis: a case report.

Danazol, an inhibitor of pituitary gonadotropin with weak androgenic effect, is currently being used for the treatment of endometriosis. In contrast to other C17 alkylated anabolic steroids, it has been rarely associated with the occurrence of primary liver tumors. We present the case of a patient with endometriosis in whom a hepatocellular adenoma was discovered after 14 years of treatment with danazol for endometriosis. The patient showed no clinical signs of a liver tumor and it was only detected by ultrasonography and magnetic resonance imaging (MRI). When long-term danazol therapy is required, regular ultrasonography for monitoring and early tumor detection is recommended.

Microscopic tumor cell dissemination in gastric cancer.

PURPOSE: There is still much controversy surrounding the prognostic significance of microscopic tumor cell dissemination in gastric cancer and its correlation with histopathologic parameters. We herein investigate such dissemination, predominantly restricted to the subserosa, in patients with gastric cancer. METHODS: Intraoperative bone marrow aspiration was done in 26 patients undergoing resection of gastric carcinoma. Peritoneal lavage could not be done in 8 of these 26 patients. The R0-resection rate was 84% (n = 22). A cytokeratin-directed antibody and an antibody directed against carcinoembryonic antigen served for the immunocytochemical detection of tumor cells, and the alkaline phosphatase antialkaline phosphatase method was used for visualization. RESULTS: The bone marrow aspirate and peritoneal lavage fluid were immunocytochemically positive in 31% and 56% of the patients, respectively. There was a trend (P = 0.10) towards higher overall survival rates in patients with negative bone marrow samples than in those with tumor cells detected in bone marrow samples. Analyzing the results of peritoneal lavage did not reveal any significant differences. In the group of T1/2 cancers, survival was significantly worse if the bone marrow was positive for tumor cells, with 3-year survival rates of 25% vs 77%, respectively (P < 0.05). CONCLUSION: The rates of tumor cell dissemination into the bone marrow or into the peritoneal cavity were within the scope of previous reports. Dissemination into the bone marrow resulted in significantly impaired survival in patients with T1 and T2 gastric carcinoma, and it did not correlate with known prognostic parameters.

Immunomodulation of tissue-engineered transplants: in vivo bone generation from methylprednisolone-stimulated chondrocytes.

Subcutaneously implanted, in vitro engineered tissue is generally affected by the immune system of the host even in autogenous transplantation. The aim of this study was to investigate immunomodulation of subcutaneously implanted tissue-engineered cartilage transplants by intramuscular methylprednisolone application. Transplants consisted of auricular rabbit chondrocytes, polylactide-polyglycolide co-polymer fleeces and species-specific fibrin or agarose. The transplants were subcutaneously implanted in the ridge. Thereafter, animals were separated into two groups, one with and one without methylprednisolone treatment. The specimens were histologically investigated after 6 and 12 weeks. Fleece fiber degradation was complete after 12 weeks, and all transplants showed areas of calcification. The corticosteroid-treated group presented pronounced trabecular bone generation without fibrous tissue infiltration. The untreated group showed sporadic islets of calcification without coherent bone formation, and adjacent fibrous tissue had infiltrated the transplants. Native controls and corticoid-treated transplants did not exhibit bone generation or signs of fibrous tissue infiltration. This study found that immunomodulation by intramuscular methylprednisolone application protects tissue-engineered autogenous chondrocyte transplants from fibrous tissue infiltration and induces trabecular bone formation.

Successful infliximab treatment of steroid and OKT3 refractory acute cellular rejection in two patients after intestinal transplantation.

Acute rejection resistant to established immunosuppressive rescue protocols remains the most prominent risk factor after intestinal transplantation. In two patients presenting with steroid-resistant severe acute cellular rejection 9 months and 2 years after intestinal transplantation, complete resolution was not achieved despite 5 and 10 days of OKT3 treatment, respectively, and high-dose triple baseline immunosuppression with tacrolimus, rapamycin, and steroids. There was a dissociated course of rejection with persistent moderate to severe rejection in the terminal portion of the graft despite complete recovery from rejection in the proximal parts. Both patients were treated with four subsequent infusions of infliximab (3 mg/kg body weight), a chimeric anti-tumor necrosis factor-alpha antibody. There was an immediate response regarding macroscopic appearance, graft histology, and clinical symptoms. Both patients recovered. In conclusion, infliximab has proven to be an effective rescue therapy in a selected group of patients with steroid and OKT3 refractory severe acute rejection after intestinal transplantation.

Late graft loss after intestinal transplantation in an adult patient as a result of necrotizing enterocolitis.

A 50-year-old recipient of an intestinal and coecal graft with sudden onset of abdominal distention and pain, lack of bowel movements, and vomiting after closure of the diagnostic ostomy 7 months after transplantation is reported. A plain abdominal radiograph revealed pneumatosis intestinalis. An angiography excluded obstruction of large vessels, however, with absent microcirculation of the intestine. Upper gastrointestinal endoscopy showed extensive ulcerative enteritis with several spontaneous perforations. The patient underwent exploration demonstrating a nonviable intestine. The entire necrotic intestine was removed. Vascular thrombosis was excluded. Clinical data, and macroscopic and histologic features of the intestinal graft were diagnostic for necrotizing enterocolitis (NEC). Though there has been evidence for the occurrence of NEC not only in premature infants but even in older infants, children and adolescents, the presented case is, to our knowledge, the first report of NEC as etiology of late graft loss after intestinal transplantation in an adult recipient.

Transforming growth factor beta 1 stimulates vascular endothelial growth factor gene transcription in human cholangiocellular carcinoma cells.

The expression pattern and functional interaction of proangiogenic factors in human cholangiocellular carcinoma (CCC) have not been fully defined. We therefore investigated the expression of vascular endothelial growth factor (VEGF) and transforming growth factor (TGF)-beta 1 as well as their respective receptors in human CCC tumor samples and further analyzed their functional interaction in vitro. Expression of VEGF, TGF-beta 1, and their receptors was examined by immunohistochemistry, in situ hybridization, quantitative competitive reverse transcription-PCR, and ELISA. VEGF promoter analysis and identification of transcription factors involved in promoter regulation were investigated using transient transfection and electrophoretic mobility shift assays. We observed strong expression of VEGF in CCC tumor cells and localization of VEGF receptors 1 and 2 in endothelial cells; in addition, coexpression of TGF-beta 1 and its receptors in tumor cells suggests a possible functional interaction between both cytokines. In vitro studies confirmed a paracrine/autocrine stimulation of VEGF by TGF-beta 1 at a transcriptional level. Additional molecular studies using 5' deletion and mutational analysis of the human VEGF promoter revealed that TGF-beta 1 stimulates VEGF through Sp1-dependent transcriptional activation. These data suggest that overexpression and functional interaction of TGF-beta 1 and VEGF might contribute to the "angiogenic switch" and the malignant phenotype in human CCC.

CMV hepatitis after liver transplantation: incidence, clinical course, and long-term follow-up.

Cytomegalovirus (CMV) hepatitis is described as the most frequent manifestation of CMV tissue invasive disease after liver transplantation. Its correlation with HLA-matching, hepatic artery thrombosis, and chronic rejection is still controversial. Risk factors, incidence, clinical course, and complications of CMV hepatitis were retrospectively analyzed in a 12-year series of 1,146 consecutive liver transplantations in 1,054 patients. All patients received only low-dose acyclovir but no gancyclovir prophylaxis. CMV infection was diagnosed by viral culture, pp65 antigenemia, or by polymerase chain reaction (PCR). CMV hepatitis was proven by liver biopsy. Treatment of CMV disease consisted of intravenous ganciclovir for a minimum of 14 days. Long-term follow-up of patients included monthly routine laboratory values and routine liver biopsies 1, 3 and 5 years after transplantation. CMV hepatitis was a rare event after liver transplantation, with a total incidence of 2.1% (24 cases). It was significantly more frequent in CMV seronegative (5.2%) than in seropositive recipients (0.7%). The leading indication in patients with CMV hepatitis was HCV cirrhosis (n = 8). The maximum number of pp65 positive white blood cells was 82 +/- 23 per 10,000 cells. Most courses manifested as isolated hepatitis; only 2 patients had disseminated disease. Nine of 24 patients had received OKT3 monoclonal antibodies because of steroid-resistant rejection before CMV hepatitis. In seronegative patients with CMV hepatitis, 71% revealed 1 or 2 HLA DR matches, in contrast to 32% in patients without CMV hepatitis. One-, 3-, and 5-year graft survival was 78%, 65%, and 59% in patients with CMV hepatitis compared with 88%, 81%, and 79% in patients without. Chronic rejection was observed in one patient, but already before onset of CMV hepatitis. Beneath D+R-constellation and OKT3 treatment as risk factors, HLA DR-matched grafts and HCV seem to favor manifestation of CMV hepatitis after liver transplantation. Long-term complications of CMV hepatitis were not observed, and especially no correlation with chronic rejection was found.

Diagnostic value of peripheral blood T-cell activation and soluble IL-2 receptor for acute rejection in liver transplantation.

BACKGROUND: T-cells play an important role in the pathogenesis of rejection. Monitoring T-cells activation markers in peripheral blood may contribute to diagnosis of acute rejection after liver transplantation (LTX). METHODS: Lymphocyte subset distribution, expression of T-cell activation markers (flow cytometry), concentration of soluble (s) interleukin-2 receptor (IL-2R) (solid phase chemiluminescence immunoassay), and liver enzymes as well as bilirubin were prospectively tested in peripheral blood samples of LTX patients with (n=69) and without acute rejection (n=50). Acute rejection was assessed by standard criteria including liver biopsies. RESULTS: Intra-individual monitoring of immune parameters revealed an up-regulation of IL-2 receptor (CD25) expression on CD4 and CD8 T-cells together with increases in sIL-2R levels in patients with acute rejections. Measuring sIL-2R levels resulted in highest diagnostic efficiency (>85%). This level of diagnostic efficiency was not reached by any other marker tested. From all conventional markers of hepatocellular integrity and function, alkaline phosphatase reached the highest level of diagnostic efficiency with 70%. CONCLUSIONS: Monitoring of up-regulation of the IL-2/IL-2R pathway represents a useful tool for assessment of acute rejection after LTX.

Overexpression of pRB in human pancreatic carcinoma cells: function in chemotherapy-induced apoptosis.

BACKGROUND: Human pancreatic adenocarcinomas are highly resistant to chemotherapy. The p16 tumor-suppressor protein is inactivated in more than 90% of human pancreatic cancers. The p16 protein transcriptionally inhibits expression of retinoblastoma tumor-suppressor gene pRB. The pRB protein transcriptionally inhibits expression of the p16 gene. Because pRB normally prevents apoptosis, we investigated whether pRB is involved in resistance to chemotherapy-induced apoptosis in pancreatic cancer cells. METHODS: pRB expression was examined by immunohistochemistry in 106 human pancreatic tissue specimens. The human pancreatic tumor cell line Capan-1 (pRB+/p16-) was stably transfected with p16 to functionally inactivate pRB. pRB gene expression was examined by western and northern blot analyses, and pRB function was assessed by electrophoretic mobility shift assays and promoter transactivation studies for the transcription factor E2F. Changes in cell sensitivity to chemotherapy were measured by assays for cytotoxicity and apoptosis. RESULTS: pRB was overexpressed in pancreatic ductal adenocarcinomas but was hardly detectable in other pancreatic malignancies, chronic pancreatitis, or nontransformed human pancreatic tissue. Expression of p16 in Capan-1 cells resulted in the loss of pRB gene and protein expression concomitant with increased activity of the transcription factor E2F, which was not detected in wild-type or control-transfected Capan-1 cells. Wild-type and control-transfected Capan-1 cells were resistant to chemotherapy-induced apoptosis, but pRB-depleted (i.e., p16-transfected) Capan-1 cells were highly sensitive. The effect was specific to pRB depletion because two other human pancreatic cancer cell lines that retained high pRB expression after p16 transfection were resistant to chemotherapy-induced apoptosis. CONCLUSIONS: Overexpression of pRB is associated with human pancreatic duct-cell cancer and may allow pancreatic cancer cells to evade chemotherapy-induced apoptosis.