Expression of GP88 (progranulin) in serum of prostate cancer patients is associated with Gleason scores and overall survival.

BACKGROUND: GP88/Progranulin is a well-recognized cell growth promoter in different cancers, and elevated serum GP88 levels have been described as negative prognostic factor in breast cancer. However, serum levels in prostate cancer (PCa) patients have not yet been studied. MATERIAL AND METHODS: We analyzed serum GP88 levels by enzyme immunosorbent assay and correlated them with clinicopathological parameters in PCa patients. PCa patients were separated into two groups based on the serum GP88 median level (low ≤44.56 ng/mL or high >44.56 ng/mL) and according to their median age (younger ≤66 years or elder patients >66 years). RESULTS: Low serum GP88 levels were more often detected in younger patients and high levels in elder patients (P=0.018; Fisher's exact test). PCa patients were separated into three groups, Gleason score (GS) ≤6; GS=7; and GS≥8. In receiver operating characteristic analyses, we could distinguish GS≤6 from GS=7 [area under the curve (AUC): 0.646; P=0.018] and GS≤6 from GS≥8 (AUC: 0.629; P=0.048) but not GS=7 from GS≥8. For survival analysis, GP88 levels were separated into two groups by an optimized cutoff value of 36.92 ng/mL. Using this GP88 stratification, all PCa patients and younger patients with a low serum GP88 level had a significantly better overall survival compared with patients with higher serum GP88 levels (log-rank test P=0.010 and P=0.024). CONCLUSION: Serum GP88 levels are significantly different depending on age and GS, and they are associated with the prognosis of PCa patients.

Serum levels of miR-320 family members are associated with clinical parameters and diagnosis in prostate cancer patients.

We studied the association of the serum levels of the microRNA family members miR-320a/-b/-c with clinico-pathological data to assess their applicability as diagnostic biomarker in prostate cancer (PCa) patients. The levels of miR-320a/-b/-c in 3 groups were evaluated by qRT-PCR (145 patients with PCa, 31 patients with benign prostatic hyperplasia (BPH) and 19 healthy controls). The levels of the three family members of miR-320 were directly correlated within each group (P < 0.001), but they differed significantly among the three groups (P < 0.001). The serum levels of the miR-320 family members were significantly increased in older patients compared to younger patients (≤ 66 years vs. > 66 years, P ≤ 0.001). In addition, the levels of all three miR-320 family members were significantly different in patients with low tumor stage compared with those with high tumor stage (miR-320a: P = 0.034; miR-320b: P = 0.006; miR-320c: P = 0.007) and in patients with low serum PSA compared with those with high serum PSA (≤ 4 ng vs. > 4 ng; miR-320a: P = 0.003; miR-320b: P = 0.003; miR-320c: P = 0.006). The levels of these miRNAs were inversely correlated with serum PSA levels. Detection in the serum samples of PCa patients with or without PSA relapse revealed higher levels of miR-320a/-b/-c in the group without PSA relapse before/after radical prostatectomy than in that with PCa relapse. In summary, the differences among the PCa/BPH/healthy control groups with respect to miR-320a/-b/-c levels in conjunction with higher levels in patients without a PSA relapse than in those with a relapse suggest the diagnostic potential of these miRNA-320 family members in PCa patients.

The combined serum levels of miR-375 and urokinase plasminogen activator receptor are suggested as diagnostic and prognostic biomarkers in prostate cancer.

This study aimed to assess the applicability of miR-375 in combination with the soluble urokinase plasminogen activator receptor (suPAR) protein as a diagnostic and/or prognostic biomarker for prostate cancer (PCa) patients. miR-375 levels by qRT-PCR and suPAR levels by ELISA were evaluated in serum samples from 146 PCa patients, 35 benign prostate hyperplasia (BPH) patients and 18 healthy controls. Antigen levels of suPAR differed between healthy controls and PCa or BPH patients, whereas miR-375 levels differed between PCa and BPH patients or healthy controls (p < 0.001). Additionally, suPAR levels differed between the Gleason sum groups GS = 7 versus GS > 7, with higher levels in the latter group (p = 0.011), and miR-375 levels were higher in the tumor stage group T3-T4 compared with the T1-T2 group (p = 0.039). A high concentration of suPAR was associated with a poor disease-specific survival (DSS; p = 0.039). The combination of suPAR and miR-375 levels identified a patient group possessing high levels for both parameters. This was associated with a poorer 10-year overall survival (OS) and DSS, with a 6.38-fold increased risk of death and a 7.68-fold increased risk of tumor-related death (p = 0.00026 and p = 0.014; univariate Cox's regression analysis). In a multivariate Cox's regression analysis PCa patients with high levels of suPAR and miR-375 showed a 5.72-fold increased risk of death in OS (p = 0.006). In summary, the differences between the PCa/BPH/healthy control cohorts for either suPAR and miR-375 levels in conjunction with the association of combined high suPAR/miR-375 levels with a poor prognosis suggest a diagnostic and prognostic impact for PCa patients.

Neutrophil gelatinase-associated lipocalin (NGAL) predicts response to neoadjuvant chemotherapy and clinical outcome in primary human breast cancer.

In our previous work we showed that NGAL, a protein involved in the regulation of proliferation and differentiation, is overexpressed in human breast cancer (BC) and predicts poor prognosis. In neoadjuvant chemotherapy (NACT) pathological complete response (pCR) is a predictor for outcome. The aim of this study was to evaluate NGAL as a predictor of response to NACT and to validate NGAL as a prognostic factor for clinical outcome in patients with primary BC. Immunohistochemistry was performed on tissue microarrays from 652 core biopsies from BC patients, who underwent NACT in the GeparTrio trial. NGAL expression and intensity was evaluated separately. NGAL was detected in 42.2% of the breast carcinomas in the cytoplasm. NGAL expression correlated with negative hormone receptor (HR) status, but not with other baseline parameters. NGAL expression did not correlate with pCR in the full population, however, NGAL expression and staining intensity were significantly associated with higher pCR rates in patients with positive HR status. In addition, strong NGAL expression correlated with higher pCR rates in node negative patients, patients with histological grade 1 or 2 tumors and a tumor size <40 mm. In univariate survival analysis, positive NGAL expression and strong staining intensity correlated with decreased disease-free survival (DFS) in the entire cohort and different subgroups, including HR positive patients. Similar correlations were found for intense staining and decreased overall survival (OS). In multivariate analysis, NGAL expression remained an independent prognostic factor for DFS. The results show that in low-risk subgroups, NGAL was found to be a predictive marker for pCR after NACT. Furthermore, NGAL could be validated as an independent prognostic factor for decreased DFS in primary human BC.

Vertebral osteoporosis: perfused animal cadaver model for testing new vertebroplastic agents.

STUDY DESIGN: Experimental study. OBJECTIVE: It was aimed to establish a cadaver model to imitate osteoporotic perfused vertebral bone and to allow for transpedicular transfer of bone cement and various new materials into vertebrae. The model was perfused to simulate vertebroplasty in the presence of transvertebral blood flow. SUMMARY OF BACKGROUND DATA: The injection of bone cement into vertebrae bears the risk of irreversible discharge of material into the venous system of the spinal canal. The bovine cadaver model studied allows visual studies of material distribution in a vertebral bone, the potential spill-out of material, and quantification of washout and disintegration phenomena. METHODS: Thoracic and lumbar vertebrae from 1-year-old calves were cut transversally into 5 mm slices, macerated, and decalcified. The softened bone slices were compressed between 2 transparent plastic discs. A standard vertebroplasty cannula (outer diameter 3.5 mm, inner diameter 2.5 mm) was inserted into the vertebral body via the pedicle to transfer the different vertebroplasty materials. Arterial blood flow was simulated by means of liquid irrigation via 2 needles in the ventral part of the vertebral body slice. Metal powder was mixed with the solution to indicate the blood flow in the bone. The model was evaluated with the vertebroplasty cement polymethylmethacrylate. RESULTS: The model permitted visualization of the insertion and distribution of vertebroplasty materials. Liquid bone cement was effused into the spinal canal as in the clinical situation. Higher modulus cement acted in the same way as in clinical vertebroplasty. Rigid vertebroplasty agents led to trabecular fractures and stable mechanical interactions with the bone and eventually moved dorsal bone fragments into the spinal canal. Sedimentation of the metal powder indicated regions of perfusion. CONCLUSION: The model simulated the clinical behavior of liquid and higher modulus vertebroplasty agents in the presence of blood flow. It enabled safe ex vivo testing of the mechanical and physical properties of alternative vertebroplasty materials under flow conditions.

Histological evidence for the existence of germ cell tumor cells showing embryonal carcinoma morphology but lacking OCT4 expression and cisplatin sensitivity.

The biological basis for manifestation of chemotherapy resistance in metastatic testicular germ cell tumors (GCT) remains obscure and is of particular clinical interest. In nonseminomatous GCT (NSGCT) the pluripotent embryonal carcinoma (EC) cells are the precursors of the manifold differentiated structures but also drive the malignant growth. They are known to be hypersensitive towards DNA-damaging agents and to express the embryonal transcription factor OCT4. We recently characterized EC cells that lack OCT4 expression and show cisplatin resistance. In the present, immunohistochemical study we analyzed the composition of NSGCT with the focus on such OCT4-negative EC cells using a NSGCT xenograft model as well as patient-derived NSGCT samples. In the xenograft model, the cisplatin-sensitive cell line H12.1 gives rise to xenografts where EC structures are mainly composed of OCT4-positive cells, whereas xenografts from the resistant cell line 1411HP exclusively comprise OCT4-negative EC areas. We found that post-chemotherapy residual metastatic tumors of patients can be comprised of exclusively OCT4-negative EC, whereas the matched testicular primary tumor harbors OCT4-positive EC. Thorough histological analyses revealed a few examples of such OCT4-negative EC cells also in the testicular primary tumor as well as in xenografts from the cisplatin-sensitive NSGCT-cell line. For these cells we propose an identity as early extraembryonal progenitor cells directly derived from OCT4-expressing EC cells. This challenges the use of the term EC cell. The data also support our hypothesis that malignant growth of resistant NSGCT may be driven by this cell type.

Prognostic impact of HIF-1alpha expression in patients with definitive radiotherapy for cervical cancer.

PURPOSE: To investigate the relationship between the hypoxia-inducible factor-(HIF-)1alpha expression in tumor tissue, tumor oxygenation and hemoglobin levels in patients with advanced cervical cancers prior to radiotherapy and the effect on clinical outcome. PATIENTS AND METHODS: The investigation included 44 patients who underwent definitive radiotherapy for advanced cervical cancers between May 1995 and March 1999. Tumor biopsies were taken prior to treatment, and HIF-1alpha expression was determined by immunohistochemistry. In the same tumor area, tumor tissue oxygenation (pO2) was measured using the Eppendorf device. RESULTS: The 5-year cancer-specific survival of all patients was 60%. Twelve of 44 tumor specimens were HIF-1alpha-negative with a significantly better 5-year survival (92 +/- 8%) versus 32 patients who were HIF-1alpha-positive (45 +/- 10%; p < 0.02). There was no correlation between HIF-1alpha expression and tumor oxygenation (p = 0.57 both for pO2 median and hypoxic fraction < 5 mmHg vs. HIF-1alpha expression). However, patients with hemoglobin levels < 11 g/dl showed elevated HIF-1alpha expression compared to patients with hemoglobin levels > 12.5 g/dl (p = 0.04). Furthermore, HIF-1alpha correlated with vascular endothelial growth factor expression (p = 0.002). In a multivariate Cox regression model, HIF-1alpha expression (relative risk [RR] = 7.5; p = 0.05) revealed an increased risk of tumor-related death. CONCLUSION: The study indicates, that endogenous tumor markers such as HIF-1alpha may serve as prognostic markers of clinical outcome concerning cervical cancer after primary radiotherapy.

Cystic fibrosis: ultrastructural changes of nasal mucosa.

Cystic fibrosis (CF) is an inherited multisystemic disorder that results in a generalized dysfunction of exocrine glands. Besides chronic obstructive pulmonary diseases, chronic sinusitis, nasal polyposis and hypertrophy of the inferior turbinates with nasal airway obstruction are typical signs. Tissue samples of the inferior turbinates and nasal polyps were taken during nasal surgery from 21 children, ranging from 3 to 16 years of age. Light- and electron microscopical examination were carried out. Furthermore, specimens of nasal mucosa of patients without chronic inflammation as controls and specimens of duodenal mucosa of patients with CF were investigated. Under a thick layer of respiratory epithelium with a high proportion of goblet cells, the seromucous glands display abnormal morphological structures with wide mucous cells and cystic dilatation. The glandular cells show inhomogeneous glandular droplets in the supranuclear cell portion. The nucleus contains dispersed chromatin as a sign of increased activity and the structures of the Golgi apparatus are clearly detectable. Apart from investigations concerning nasal polyps in CF, studies on the different morphological changes of nasal mucosa at the electron microscopic level are rare. This histological study focuses on various morphological changes of nasal glands at the ultrastructural level in correlation with typical symptoms in CF. In addition, a comparison with electron microscopic findings of CF-enteropathies is proposed. These findings could help to bring information concerning new morphological aspects in the pathophysiology of patients with CF.

Reliability and validity of needle biopsy evaluation of breast-abnormalities using the B-categorization--design and objectives of the Diagnosis Optimisation Study (DIOS).

BACKGROUND: The planned nationwide implementation of mammography screening 2007 in Germany will increase the occurrence of mammographically detected breast abnormalities. These abnormalities are normally evaluated by minimal invasive core biopsy. To minimize false positive and false negative histological findings, quality assurance of the pathological evaluation of the biopsies is essential. Various guidelines for quality assurance in breast cancer diagnosis recommend applying the B-classification for histopathological categorization. However, to date there are only few studies that reported results about reliability and validity of B-classification. Therefore, objectives of our study are to determine the inter- and intraobserver variability (reliability study) and construct and predictive validity (validity study) of core biopsy evaluation of breast abnormalities. This paper describes the design and objectives of the DIOS Study. METHODS/DESIGN: All consecutive asymptomatic and symptomatic women with breast imaging abnormalities who are referred to the University Hospital of Halle for core breast biopsy over a period of 24 months are eligible. According to the sample size calculation we need 800 women for the study. All patients in the study population underwent clinical and radiological examination. Core biopsy is performed by stereotactic-, ultrasound- or magnetic resonance (MR) guided automated gun method or vacuum assisted method. The histopathologic agreement (intra- and interobserver) of pathologists and the histopathologic validity will be evaluated. Two reference standards are implemented, a reference pathologist and in case of suspicious or malignant findings the histopathologic result of excision biopsy. Furthermore, a self administrated questionnaire which contains questions about potential risk factors of breast cancer, is sent to the participants approximately two weeks after core biopsy. This enables us to run a case-control-analysis (woman with breast cancer histological verified after excision are defined as cases, woman without malignant breast lesions are defined as controls) to investigate the predictive values of various risk factors on breast cancer risk. CONCLUSION: The analysis of reliability and validity of the histopathological evaluation of core biopsy specimens of breast abnormalities is intended to provide important information needed for a high quality in breast cancer diagnostic and for planning of treatment strategies.

Invasive aspergillosis of the paranasal sinuses and the skull base.

Invasive aspergillosis (IA) originating from the paranasal sinuses can cause an intracranial growth mainly along the skull base and larger vessels. This study reports our experience in the diagnosis and treatment of a series of patients with IA. A retrospective chart review of four patients with chronic invasive intracranial aspergillosis was performed. Clinical signs, physical examinations, radiographs, histological samples, and outcome were demonstrated. The patients demonstrated different symptoms like exophthalmus, ophthalmoplegia, loss of vision, and hypaesthesia of the ophthalmic and maxillary nerve. Computed tomography and MRI revealed extensive sino-orbital and skull base lesions. The patients were treated with aggressive endonasal debridement, intravenous antifungal agents and daily irrigations with antimycotic suspensions. Furthermore, we applied hyperbaric oxygenation. Two patients died from complications due to subarachnoidal hemorrhage and accompanied complications respectively. Despite the high mortality rate patients with an invasive aspergillosis can be effectively treated in some cases by an early and rigorous treatment schedule using all surgical and conservative therapeutic options.

Rhinitis medicamentosa: electron microscopic changes of human nasal mucosa.

OBJECTIVE: Prolonged application of nasal vasoconstrictors causes rhinitis medicamentosa (RM). Nasal obstruction is induced by rebound swelling when the decongestive effect has disappeared. The aim of this study was to demonstrate ultrastructural changes in RM. STUDY DESIGN AND SETTING: Tissue samples of inferior turbinates from 22 patients with RM and 10 patients without rhinitis were taken during nasal surgery. Ultrathin sections were investigated by using a transmission electron microscope (TEM). RESULTS: The TEM findings revealed severe epithelial damages such as loss of ciliated cells. In the subepithelial region, the vascular endothelium showed gaps and ruptures of basal lamina. CONCLUSION: RM is a drug-induced damage of human nasal mucosa. Loss and destruction of ciliated epithelial cells are the morphologic correlation of the disturbed mucociliary clearance. In addition, vascular endothelium revealed ultrastructural changes. This could be caused by an increased vascular permeability with consecutive interstitial edema. SIGNIFICANCE: This study demonstrated new morphological aspects of rhinitis medicamentosa.

INSL3 in the benign hyperplastic and neoplastic human prostate gland.

The human prostate gland is a well known source of H1 and H2 relaxin but information is lacking on the expression and potential role of the INSL3 peptide hormone within the prostate gland. In the present study we have investigated the expression of human INSL3 in patients with benign prostate hyperplasia (BPH), prostate intraepithelial neoplasia (PIN) and prostate carcinoma tissues. Of the prostate epithelial cells, strongest INSL3 expression was detected in the basal epithelial cell compartment. Weaker INSL3 mRNA expression and immunoreactive INSL3 production were observed in secretory epithelial cells and in interstitial smooth muscle cells. Prostate epithelial cells were also a source for transcripts encoding the INSL3 receptor LGR8 suggesting the presence of an autocrine/paracrine INSL3-LGR8 ligand-receptor system within the human prostate. Three human prostate carcinoma cell lines displayed differential gene activity for INSL3 and LGR8. While LNCaP was devoid of INSL3, the androgen-insensitive PC-3 and the stromal prostate cell line hPCP co-expressed INSL3 and LGR8 transcripts. In addition to expressing INSL3 mRNA, the LGR8-negative DU-145 also expressed an INSL3 splice form formerly demonstrated in thyroid carcinoma cells. When incubated with recombinant INSL3, PC-3 cells showed significantly increased intracellular cAMP levels indicating functional LGR8 receptors. INSL3 did not alter the proliferative or metabolic activity of PC-3 carcinoma cells. Instead, PC-3 responded to INSL3 with significantly enhanced tumor cell motility and a transcriptional down-regulation of ErbB receptors and EGF. All-trans-retinoic acid was demonstrated in PC-3 to up-regulate LGR8 gene activity in a dose- and time-dependent manner while having no effect on INSL3 gene activity. In conclusion, we have identified a functional INSL3-LGR8 ligand-receptor system in human prostate carcinoma cells.