Bone mineralisation adjacent to cemented and uncemented acetabular cups: analysis by [18F]-fluoride-PET in a randomised clinical trial.

PURPOSE: We present a randomised clinical trial using F-PET/CT to analyse new bone metabolic mineralisation adjacent to acetabular cups following total hip arthoplasty (THA). PATIENTS AND METHODS: THA was performed on 26 patients (26 cases) with hip OA. Patients with hip osteoarthritis (OA) were randomly assigned to operations with cemented or uncemented acetabular components. The contralateral, healthy acetabulum was used as referent for normal bone metabolism. The patients were analysed with radiography, clinical scoring, and F-PET/CT preoperatively, and at 6 weeks and 6 months postoperatively. RESULTS: No major complications were recorded, and clinical results were good in all patients. Radiography showed all cups to be stable. The bone-forming activity, as measured by F-PET/CT, was quantified as standardised uptake values (SUV). The mean SUV was 4.6 (6 weeks) and 3.5 (6 months) around the uncemented cups, and 4.8 and 4.0, respectively, for the cemented cups. Normal healthy bone metabolism in the referent was 2.8 and 2.7 SUV at 6 weeks and 6 months, respectively. P < 0.01 for the cemented group at 6 weeks and 6 months, for the uncemented group only at 6 weeks. INTERPRETATION: An acetabulum affected by OA has elevated SUV activity. Both cemented and uncemented cups had elevated bone metabolic activity at 6 weeks. The raised activity was interpreted as an effect from bone mineralisation secondary to surgical trauma and healing, and to the OA. At 6 months, activity was more normalised for the uncemented group than for the cemented, suggesting healing may terminate faster in the uncemented group. Postoperative bone metabolic activity can be analysed in detail by F-PET/CT.ClinicalTrials.gov Identifier: NCT01623687.

Fingerprint pattern of bone mineralisation on cemented and uncemented femoral stems: analysis by [18F]-fluoride-PET in a randomised clinical trial.

PURPOSE: We present a randomised clinical study using 18F-fluoride positron emission tomography/computed tomography (F-PET/CT) to analyse the osteoblastic part of bone metabolism (new bone mineralisation) in periprosthetic bone adjacent to femoral stems following total hip arthoplasty (THA) surgery. Patients with hip osteoarthritis were randomly assigned to THA surgery with cemented or uncemented femoral components. PATIENTS AND METHODS: THA was performed on 26 patients (26 cases) with hip osteoarthritis. The patients received either an uncemented HA-coated femoral stem or a cemented one. The contralateral healthy femur was used as referent for normal bone metabolism. The patients were analysed with clinical score, radiography and F-PET/CT preoperatively, and postoperatively at 6 weeks and 6 months. After 2 years, clinical score and radiography was analysed again. We used the Polar Map system for analysing and presenting the PET results in 13 regions of interest adjacent to the whole stem. RESULTS: The clinical results were good in all patients; there were no major complications. Radiographically, all stems were stable. PET analyses after 6 weeks showed that bone mineralising activity was significantly higher around the uncemented stems, both compared to the cemented group and to the contralateral healthy reference femur group. The cemented group also had elevated activity but only at a barely significant level. INTERPRETATION: Mineralising activity analysed with F-PET/CT was significantly higher for the uncemented group and also decreased at a slower rate. F-PET/CT is a useful new tool for analysing secondary stabilisation of femoral stems after THA. The study was registered at ClinicalTrials.gov (identifier NCT01623687).

miR-203 and miR-320 Regulate Bone Morphogenetic Protein-2-Induced Osteoblast Differentiation by Targeting Distal-Less Homeobox 5 (Dlx5).

MicroRNAs (miRNAs) are a family of small, non-coding RNAs (17-24 nucleotides), which regulate gene expression either by the degradation of the target mRNAs or inhibiting the translation of genes. Recent studies have indicated that miRNA plays an important role in regulating osteoblast differentiation. In this study, we identified miR-203 and miR-320b as important miRNAs modulating osteoblast differentiation. We identified Dlx5 as potential common target by prediction algorithms and confirmed this by knock-down and over expression of the miRNAs and assessing Dlx5 at mRNA and protein levels and specificity was verified by luciferase reporter assays. We examined the effect of miR-203 and miR-320b on osteoblast differentiation by transfecting with pre- and anti-miRs. Over-expression of miR-203 and miR-320b inhibited osteoblast differentiation, whereas inhibition of miR-203 and miR-320b stimulated alkaline phosphatase activity and matrix mineralization. We show that miR-203 and miR-320b negatively regulate BMP-2-induced osteoblast differentiation by suppressing Dlx5, which in turn suppresses the downstream osteogenic master transcription factor Runx2 and Osx and together they suppress osteoblast differentiation. Taken together, we propose a role for miR-203 and miR-320b in modulating bone metabolism.

Second generation sequencing of microRNA in Human Bone Cells treated with Parathyroid Hormone or Dexamethasone.

We investigated the impact of treatment with parathyroid hormone (PTH) and dexamethasone (DEX) for 2 and 24h by RNA sequencing of miRNAs in primary human bone (HOB) cells. A total of 207 million reads were obtained, and normalized absolute expression retrieved for 373 most abundant miRNAs. In naïve control cells, 7 miRNAs were differentially expressed (FDR<0.05) between the two time points. Ten miRNAs exhibited differential expression (FDR <0.05) across two time points and treatments after adjusting for expression in controls and were selected for downstream analyses. Results show significant effects on miRNA expression when comparing PTH with DEX at 2h with even more pronounced effects at 24h. Interestingly, several miRNAs exhibiting differences in expression are predicted to target genes involved in bone metabolism e.g. miR-30c2, miR-203 and miR-205 targeting RUNX2, and miR-320 targeting ß-catenin (CTNNB1) mRNA expression. CTNNB1and RUNX2 levels were decreased after DEX treatment and increased after PTH treatment. Our analysis also identified 2 putative novel miRNAs in PTH and DEX treated cells at 24h. RNA sequencing showed that PTH and DEX treatment affect miRNA expression in HOB cells and that regulated miRNAs in turn are correlated with expression levels of key genes involved in bone metabolism.

Podocalyxin as a Prognostic Marker in Gastric Cancer.

BACKGROUND: Podocalyxin-like 1 (PODXL) is a cell-adhesion glycoprotein associated with aggressive tumor phenotype and poor prognosis in several forms of cancer. The aim of this study was to investigate PODXL expression in gastric cancer by use of two different antibodies. METHODS: By tumor-tissue microarrays and immunohistochemistry we evaluated PODXL expression in tumor specimens from 337 patients who underwent surgery for gastric adenocarcinoma at Helsinki University Hospital. We used two different antibodies: HPA2110, which is a polyclonal antibody and an in-house monoclonal antibody called HES9, to investigate the association of PODXL expression with clinicopathologic variables and patient survival. RESULTS: PODXL staining was positive by the polyclonal antibody in 153 (57.5%) cases and by the monoclonal antibody in 212 (76%). Polyclonal antibody expression was associated with intestinal cancer type (p<0.001). Monoclonal antibody staining was associated with age over 66 (p = 0.001), with intestinal cancer (p<0.001), and with small tumor size (≤ 5 cm; p = 0.024). Both antibodies were associated with high S-phase fraction (p = 0.022; p = 0.010), and high tumor proliferation index (Ki-67; p = 0.003; p = 0.001). PODXL positivity by the polyclonal antibody indicated reduced gastric-cancer-specific 5-year survival of 24.0% (95% CI 16.9-31.1), compared to 43.3% (95% CI 33.7-52.9) for patients with PODXL negativity (p = 0.001). The result remained significant in multivariable analysis (HR = 3.17; 95% CI 1.37-7.34, p = 0.007). CONCLUSION: In gastric cancer, PODXL expression by the polyclonal antibody HPA2110 is an independent marker of poor prognosis.

Expression of Stem Cell-associated Marker HES77 in Rectal Neuroendocrine Tumors.

BACKGROUND: Expression of novel stem cell-associated marker human embryonic stem cell 77 (HES77) was studied in rectal neuroendocrine tumors (NETs), which comprise 10 to 15% of gastroenteropancreatic NETs, some with metastatic potential. MATERIALS AND METHODS: WHO 2010 classification was applied, and immunohistochemical positivity for HES77 was assessed in 72 primary tumors and 6 metastases. Correlations were calculated between HES77 expression, metastasis and patient survival. RESULTS: Expression of HES77 strongly positively correlated with metastatic potential and poorer prognosis. The proliferative index determined in the metastasis did not correlate with patient survival. CONCLUSION: Novel stem cell-associated marker HES77 has a strong prognostic value in patients with rectal NETs and may be useful in selecting those who are at-risk for developing metastatic disease, and who may benefit from intensive adjuvant therapy. Proliferative index in the metastasis did not predict for outcome. Characterization of the HES77 epitope would certainly enhance the interest in the antibody.

Global miRNA expression and correlation with mRNA levels in primary human bone cells.

MicroRNAs (miRNAs) are important post-transcriptional regulators that have recently introduced an additional level of intricacy to our understanding of gene regulation. The aim of this study was to investigate miRNA-mRNA interactions that may be relevant for bone metabolism by assessing correlations and interindividual variability in miRNA levels as well as global correlations between miRNA and mRNA levels in a large cohort of primary human osteoblasts (HOBs) obtained during orthopedic surgery in otherwise healthy individuals. We identified differential expression (DE) of 24 miRNAs, and found 9 miRNAs exhibiting DE between males and females. We identified hsa-miR-29b, hsa-miR-30c2, and hsa-miR-125b and their target genes as important modulators of bone metabolism. Further, we used an integrated analysis of global miRNA-mRNA correlations, mRNA-expression profiling, DE, bioinformatics analysis, and functional studies to identify novel target genes for miRNAs with the potential to regulate osteoblast differentiation and extracellular matrix production. Functional studies by overexpression and knockdown of miRNAs showed that, the differentially expressed miRNAs hsa-miR-29b, hsa-miR-30c2, and hsa-miR-125b target genes highly relevant to bone metabolism, e.g., collagen, type I, α1 (COL1A1), osteonectin (SPARC), Runt-related transcription factor 2 (RUNX2), osteocalcin (BGLAP), and frizzled-related protein (FRZB). These miRNAs orchestrate the activities of key regulators of osteoblast differentiation and extracellular matrix proteins by their convergent action on target genes and pathways to control the skeletal gene expression.

Podocalyxin Is a Marker of Poor Prognosis in Pancreatic Ductal Adenocarcinoma.

AIM OF THE STUDY: Podocalyxin-like 1 is a transmembrane glyco-protein whose overexpression associates in many cancers with poor prognosis and unfavorable clinicopathological characteristics. Until now, its prognostic value has never been studied in pancreatic ductal adenocarcinoma (PDAC). The aim of this study was to investigate podocalyxin expression in PDAC by a novel monoclonal antibody and a commercially available polyclonal antibody. PATIENTS AND MATERIALS: With tissue microarrays and immuno-histochemistry, podocalyxin expression evaluation involved 168 PDAC patients. The associations of the podocalyxin tumor expression with clinicopathological variables were explored by Fisher's exact test and the linear-by-linear test. Survival analyses were by Kaplan-Meier analysis and the Cox proportional hazard model. RESULTS: The polyclonal antibody revealed membranous podocalyxin expression in 73 (44.0%) specimens and the monoclonal antibody was highly expressed in 36 (21.8%) cases. Membranous expression by the polyclonal antibody was associated with T classification (p=0.045) and perineural invasion (p=0.005), and high expression by the mono-clonal antibody with poor differentiation (p=0.033). High podocalyxin expression associated significantly with higher risk of death from PDAC by both the polyclonal antibody (hazard ratio (HR) = 1.62; 95% confidence interval (CI) 1.12-2.33; p=0.01) and the monoclonal antibody (HR = 2.10, 95% CI 1.38-3.20; p<0.001). The results remained significant in multivariate analysis, adjusted for age, gender, stage, lymph node ratio (≥/< 20%), and perivascular invasion (respectively as HR = 2.03; 95% CI 1.32-3.13, p=0.001; and as HR = 2.36; 95% CI 1.47-3.80, p<0.001). CONCLUSION: We found podocalyxin to be an independent factor for poor prognosis in PDAC. To our knowledge, this is the first such report of its prognostic value.

A novel stem cell associated marker identified by monoclonal antibody HESC5:3 differentiates between neoplastic lesions in follicular thyroid neoplasms.

Follicular thyroid lesions are the bane of cytopathology. Differentiation between adenoma and carcinoma is impossible, and often these neoplasms are indistinguishable even from uninodular goitre. In other cancers as well, a theory of stem cells as the origin of cancer has been discussed in thyroid carcinogenesis. We aimed to examine a novel stem cell associated marker identified by monoclonal antibody HESC5:3 in follicular lesions in an attempt to find a marker for differential diagnosis in thyroid cytopathology. HESC5:3 was raised against and is specific for undifferentiated human embryonic stem cells. The epitope of this novel antibody is to be defined. Immunohistochemical expression of HESC5:3 was examined in clinical material comprised of follicular neoplasms (83 adenomas, 43 carcinomas) and non-neoplastic lesions (41 goitrous, 22 hyperplastic, 23 normal tissue specimens). Staining differed significantly between neoplastic and non-neoplastic lesions. Nuclear staining was increased in non-neoplastic cells, whereas in neoplastic cells expression was mainly cytoplasmic. There was no difference between benign and malignant lesions, suggesting a role in early tumourigenesis. In conclusion, the HESC5:3 epitope may be of benefit as a neoplasia marker in distinguishing between uninodular goitre and neoplasia. Characterization of the epitope would increase the interest in this promising new stem cell associated marker.

Legg-Calvé-Perthes disease and the risk of ADHD, depression, and mortality.

BACKGROUND AND PURPOSE: Hyperactive behavior pattern (such as attention deficit hyperactivity disorder (ADHD)) is proposed to be present in individuals with Legg-Calvé-Perthes disease (LCPD). We investigated whether individuals with LCPD have a higher risk of ADHD, depression, and mortality. SUBJECTS AND METHODS: We identified 4,057 individuals with LCPD in Sweden during the period 1964-2011. 40,570 individuals without LCPD were randomly selected from the Swedish general population and matched by year of birth, sex, and region (control group). We used Cox proportional hazard regression to estimate the relative risks. RESULTS: Compared to the control group, individuals with LCPD had a raised hazard ratio (HR) of 1.5 (95% CI: 1.2-1.9) for ADHD. The risks were higher for female individuals (HR = 2.1, CI: 1.3-3.5) than for male individuals (HR = 1.4, CI: 1.1-1.8). Individuals with LCPD had a modestly higher hazard ratio for depression (HR = 1.3, CI: 1.1-1.5) than the control group. Furthermore, individuals with LCPD had a slightly higher mortality risk than the control group (HR = 1.2, CI: 1.0-1.4) INTERPRETATION: Individuals with LCPD have a higher risk of ADHD. Hyperactivity could expose the femoral head to higher mechanical stress and contribute to the etiology of LCPD. The higher risk of depression may be due to the burden of LCPD itself or could reflect neurobehavioral aspects of ADHD changing into depression later in life. Individuals with LCPD have a higher mortality risk, with higher risk of suicide and cardiovascular diseases.

Podocalyxin is a marker of poor prognosis in colorectal cancer.

BACKGROUND: Over two decades ago, a proposal was that two different colorectal cancer (CRC) entities existed, based on tumour location either proximal (right) or distal (left) of the splenic flexure. Proximal and distal tumours exhibit different clinical, epidemiological, and biological characteristics. Improvement of the prognostic evaluation of CRC requires new molecular markers. Podocalyxin-like 1 (PODXL), an anti-adhesive transmembrane sialomucin, is associated with an aggressive tumour phenotype and poor prognosis. For colorectal cancer, it has been suggested to be a marker of poor prognosis. The aim of this study was to investigate the role of PODXL in CRC by use of a novel monoclonal antibody. METHODS: In 1983-2001, 840 consecutive colorectal cancer patients were treated at Helsinki University Central Hospital, of whom 767 were successfully scored for PODXL immunohistochemical expression from tumour tissue microarrays by use of a novel monoclonal in-house antibody. Associations of PODXL expression and tumour location with other clinicopathological variables were explored by Fisher's exact-test, linear-by- linear association test, and binary logistic regression. Survival analyses were done by the Kaplan-Meier method and Cox proportional hazards model. RESULTS: PODXL protein expression was high in 44 (5.7%) specimens. High expression associated strongly with poor differentiation (p < 0.0001), advanced stage (p = 0.011), and location of the tumour in the right hemicolon (RHC) (p < 0.001). Tumours of the RHC were more poorly differentiated (p < 0.0001) and showed higher PODXL expression (p < 0.001).High PODXL expression associated significantly with higher risk for disease-specific death from CRC (hazard ratio (HR) = 2.00; 95% confidence interval (CI) 1.31-3.06, p = 0.001) and also in the subgroups of left hemicolon (LHC) cancers (HR = 2.60; 95% CI 1.45-4.66, p = 0.001) and rectal cancers (HR = 3.03; 95% CI 1.54-5.60, p = 0.001). Results remained significant in multivariable analysis (respectively, HR = 1.82; 95% CI 1.15-2.86, p = 0.01; HR = 2.59; 95% CI 1.41-4.88, p = 0.002; and HR = 2.69; 95% CI 1.30-5.54, p = 0.007). CONCLUSION: Podocalyxin was an independent factor for poor prognosis in colorectal cancer and in the subgroups of left hemicolon and rectum. This is, to our knowledge, the first evidence of such difference in PODXL expression, its function possibly being dependent upon tumour location.

A comparative study of two PODXL antibodies in 840 colorectal cancer patients.

BACKGROUND: Podocalyxin (PODXL) is a transmembrane sialomucin, whose aberrant expression and/or allelic variation associates with poor prognosis and unfavourable clinicopathological characteristics in different cancers. Membranous expression of PODXL has been suggested to be an independent marker of poor prognosis in colorectal cancer (CRC), and previously by an in-house monoclonal antibody, we showed that also cytoplasmic overexpression of PODXL predicts poor prognosis. The aim of this study was to compare two PODXL antibodies with different epitopes case-by-case in CRC patients. METHODS: Of 840 consecutively operated CRC patients from Helsinki University Central Hospital, PODXL expression by polyclonal HPA 2110 antibody was evaluated from 780. Associations of PODXL expression with clinicopathological parameters and the impact of PODXL expression on survival were assessed. Kappa-value was used to assess the comparability of the two antibodies. RESULTS: Membranous PODXL expression associated with unfavourable clinicopathological parameters and with higher risk for disease-specific death from CRC within 5 years (unadjusted hazard ratio (HR) = 1.90; 95% confidence interval (CI) (1.32-2.75); adjusted HR = 1.64; 95% CI (1.11-2.43)). The comparability of expressions by the two antibodies was low (kappa =0.219, standard error 0.060, p < 0.0001). Combination of two antibodies identified a group of patients with even worse prognosis (unadjusted HR = 6.00; 95% CI (3.27-13.0); adjusted HR = 2.14; 95% CI (1.12-4.07)). CONCLUSION: Membranous expression by the polyclonal PODXL antibody and cytoplasmic overexpression by the monocolonal PODXL antibody are both independent markers of poor prognosis, but they recognise different groups of patients, both of which have poor prognosis. The combined use of the antibodies reveals a group with an even worse prognosis. The biological reasons for the difference between antibodies warrant further studies.

Comparison of reverse transcription quantitative real-time PCR, flow cytometry, and immunohistochemistry for detection of monoclonality in lymphomas.

In healthy humans, 60-70% of the B lymphocytes produce kappa light chains, while the remaining cells produce lambda light chains. Malignant transformation and clonal expansion of B lymphocytes lead to an altered kappa : lambda expression ratio, which is an important diagnostic criteria of lymphomas. Here, we compared three methods for clonality determination of suspected B cell lymphomas. Tumor biopsies from 55 patients with B cell malignancies, 5 B-lymphoid tumor cell lines, and 20 biopsies from patients with lymphadenitis were analyzed by immunohistochemistry, flow cytometry, and reverse transcription quantitative real-time PCR. Clonality was determined by immunohistochemistry in 52/53 cases, flow cytometry in 30/39 cases, and reverse transcription quantitative real-time PCR in 33/55 cases. In conclusion, immunohistochemistry was superior to flow cytometry and reverse transcription quantitative real-time PCR for clonality identification. Flow cytometry and reverse transcription quantitative real-time PCR analysis has complementary values. In a considerable number of cases tumor cells produced both kappa and lambda light chain transcripts, but only one type of light chain peptide was produced.

Analysis of bone mineralization on uncemented femoral stems by [18F]-fluoride-PET: a randomized clinical study of 16 hips in 8 patients.

PURPOSE: We present the first study using fluoride-positron emission CT (F-PET/CT) to analyze mineralization of bone in the femur adjacent to uncemented stems following total hip arthroplasty (THA). We studied patients who were operated bilaterally for osteoarthritis with 2 different stems during the same surgical session. PATIENTS AND METHODS: THA was performed bilaterally during the same surgical session in 8 patients with bilateral osteoarthritis of the hip. An SL-PLUS stem was inserted in one hip and a BetaCone stem was inserted in the contralateral hip, with randomization of side and sequence. A second group of 12 individuals with a normal healthy hip was used as reference for normal bone metabolism. Clinical and radiographic evaluation was performed preoperatively, postoperatively, and at 2 years. We used [18F]-fluoride-PET/CT to analyze bone mineralization adjacent to the stems 1 week, 4 months, and 12 months after surgery. We modified the Polar Map system to fit the upper femur for analysis and presentation of the PET results from 12 regions of interest adjacent to the whole stem. RESULTS: The clinical results were good at 2 years. By radiography, all stems were stable. At PET analyses 1 week after surgery, the activity was higher for the SL-PLUS group than for the BetaCone group. The activity was statistically significantly higher for both stems than the reference values at 4 months, and was most pronounced in the upper femur. At one year, the activity had declined more for the BC group than for the SL group. INTERPRETATION: The bone mineralization activity varied between different regions for the same stem and between different time periods for each group. F-PET/CT is a novel and valuable tool for analysis of bone mineralization patterns around uncemented femoral stems in detail. The combination of PET/CT analysis and the modified Polar Map system may provide a useful tool for future studies of metabolic bone responses to prosthetic implants.

Rats treated with AZD2858, a GSK3 inhibitor, heal fractures rapidly without endochondral bone formation.

Fracture healing is a complex interplay between endochondral and intramembranous bone formation processes. The canonical Wnt/ß-catenin pathway enhances new bone formation and may play a role in fracture healing. Glycogen synthase kinase 3ß (GSK3ß) is a key regulator of ß-catenin degradation. In this study, we investigate the effects of AZD2858, an orally bioactive GSK3 inhibitor, on fracture healing. Femoral fractures were produced in rats after the insertion of a femoral nail. The rats were treated with oral administration of AZD2858 at a dose of 30 µmol/kg (20mg/kg) daily for up to 3 weeks, while control animals were administered vehicle. At 4days, and at 1, 2 and 3 weeks, histological analysis was performed, and at the 2 and 3 week time points, we performed peripheral quantitative computed tomography (pQCT), X-rays, and four-point bending tests. Peripheral QCT showed an increase in both mineral density (of 28% at 2 weeks and 38% at 3weeks) and mineral content (of 81% at 2 weeks and 93% at 3 weeks) in the calluses from AZD2858 treated animals as compared to vehicle treated animals. Histological analysis demonstrated that rats treated with GSK3 inhibitor healed their fractures rapidly, but without the pre-formation of cartilage tissue. Furthermore, four-point bending tests of fractured femora from animals treated for 2 and 3 weeks showed an increase in strength in treated animals compared to their vehicle-treated controls. In conclusion, AZD2858, a potent GSK3 inhibitor, has a substantial impact on fracture healing. The fractures healed with a bony callus without an obvious endochondral component, suggesting that AZD2858 drives mesenchymal cells into the osteoblastic pathway. This leads to direct bone repair in an unstable fracture milieu.

Suitability of quality control materials for prostate-specific antigen (PSA) measurement: inter-method variability of common tumor marker control materials.

BACKGROUND: Quality control materials with minimal inter-assay differences and clinically relevant proportions of different molecular forms of the analyte are needed to optimize intra- and inter-laboratory accuracy and precision. METHODS: We assessed if clinically relevant total prostate-specific antigen (tPSA) levels were present in seven commercially available Multi Constituent Tumor Marker Controls (MC-TMC). Further, we determined the concentration of free PSA (fPSA) and calculated the percentage of free PSA (%fPSA) in all materials. Finally, we determined variability of TMC materials across several commonly used PSA platforms. RESULTS: All MC-TMC materials contained at least one concentration of tPSA in normal and pathologic range. Control materials varied in the amount of fPSA and %fPSA, with most controls consisting of fPSA only and only one MC-TMC containing medically relevant levels of around 35% fPSA. Only a minority of MC-TMC materials showed minimal variability across four PSA methods while the majority of PSA controls showed wide inter-method differences. CONCLUSIONS: Use of many commercially available controls for PSA could lead to biased PSA measurements because they contain medically irrelevant proportions of fPSA and show significant variation among different PSA assay platforms.

Analysis of bone formation on porous and calcium phosphate-coated acetabular cups: a randomised clinical [18F]fluoride PET study.

We present a study using Fluoride-Positron Emission Tomography (F-PET/CT) to analyse new bone formation in periacetabular bone adjacent to press fit cups following THA. In 16 THA (8 patients) with bilateral hip osteoarthritis simultaneous bilateral total hip arthroplasty (THA) was performed, employing electrochemically applied calcium phosphate coated (HA) cups or porous-coated (PC) cups allocated at random to compare the two sides. A reference group of 13 individuals with a normal healthy hip was used to determine 'normal' bone metabolism. [18F]fluoride -PET/CT was used to analyze bone formation adjacent to the cups 1 week, 4 months and 12 months after surgery. Clinical and radiographic evaluation was performed preoperatively, postoperatively and at 2 years. Bone forming activity had a mean of 5.71, 4.69 and 3.47 SUV around the HA- and 5.04, 4.80 and 3.50 SUV around the PC-cups at 1 week, 4 months and 12 months respectively. Normal bone metabolism was 3.68 SUV. After 1 year activity had declined to normal levels for both groups. The clinical results were good in all cases. HA coating resulted in higher uptake indicating higher bone forming activity after 1 week. F-PET/CT is a valuable tool to analyse bone formation and secondary stabilisation of an acetabular cup.

Targeting free prostate-specific antigen for in vivo imaging of prostate cancer using a monoclonal antibody specific for unique epitopes accessible on free prostate-specific antigen alone.

This study investigated the feasibility of targeting the free, unbound forms of prostate-specific antigen (fPSA) for in vivo imaging of prostate adenocarcinomas (PCa), as PSA is produced and secreted at abundance during every clinical stage and grade of PCa, including castration-resistant disease. We injected (125)I-labeled monoclonal antibody PSA30 (specific for an epitope uniquely accessible on fPSA alone) intravenously in male nude mice carrying subcutaneous xenografts of LNCaP tumors (n=36). Mice were sacrificed over a time course from 4 hours to 13 days after injecting (125)I-labeled PSA30. Tissue uptake of (125)I-PSA30 at 48 and 168 hours after intravenous injection was compared with two clinically used positron emission tomography radiopharmaceuticals, (18)F-fluoro-deoxy-glucose ((18)F-FDG) or (18)F-choline, in cryosections using Digital AutoRadiography (DAR) and also compared with immunohistochemical staining of PSA and histopathology. On DAR, the areas with high (125)I-PSA30 uptake corresponded mainly to morphologically intact and PSA-producing LNCaP cells, but did not associate with the areas of high uptake of either (18)F-FDG or (18)F-choline. Biodistribution of (125)I-PSA30 measured in dissected organs ex vivo during 4 to 312 hours after intravenous injection demonstrated maximum selective tumor uptake 24-48 hours after antibody injection. Our data showed selective uptake in vivo of a monoclonal antibody highly specific for fPSA in LNCaP cells. Hence, in vivo imaging of fPSA may be feasible with putative usefulness in disseminated PCa.

Detection of human papillomavirus oncoprotein E7 in liquid-based cytology.

The selection and characterization of a set of mouse mAbs against high-risk human papillomavirus (HPV) E7 oncoprotein and the development of protocols for immunocytochemistry (ICC) are described here. A large number of antibodies raised towards HPV16 and 18 E7 were tested for high-risk specificity by ELISA using a panel of HPV E7 proteins. Antibodies detecting low-risk E7 were discarded, resulting in 38 high-risk HPV E7-specific antibodies. The corresponding epitopes were mapped using overlapping HPV E7 fragments displayed on phage particles. Functionality in ICC against formalin-fixed cervical cancer cell lines was demonstrated for ten mAbs; their high-risk specificity was confirmed by Western blot analysis and ICC on transiently transformed cells expressing high- or low-risk HPV E7. These mAbs were specific for one or several of the high-risk strains HPV16, 18, 31, 35 and 45. Specific E7 staining of liquid-based cytology (LBC) samples was demonstrated for seven mAbs and optimized protocols were established. The E716-41 and E718-79 mAbs demonstrated particularly strong and specific staining of cells stored in LBC fluid for at least 6 months. It is proposed that the high-risk HPV E7 staining protocols established in this study may have the potential to be included in a complementary test for the detection and identification of malignantly transformed cells, in for example atypical squamous cells of undetermined significance samples.

Metabolic development of necrotic bone in the femoral head following resurfacing arthroplasty. A clinical [18F]fluoride-PET study in 11 asymptomatic hips.

BACKGROUND AND PURPOSE: One concern regarding resurfacing arthroplasty is the viability of the diminished femoral head and the postoperative risk of collapse, or a femoral neck fracture. (18)F-fluoride positron emission tomography (F-PET) enables us to assess bone viability despite there being a covering metal component. By F-PET studies, we recently showed the absence of metabolism in the remaining part of femoral heads, 1-4 years after surgery in 11 of 46 consecutive cases. We now present the further development of bone metabolism in these 11 cases. PATIENTS AND METHODS: 10 patients (11 chips) with previously shown loss of femoral head metabolism were evaluated by radiography and repeated F-PET scans, 3-6.5 years after surgery. The size of the area with low (18)F-fluoride PET uptake in the femoral head was compared to that in earlier PET images. RESULTS: No patients had any clinical symptoms; nor was any necrotic bone area visible in plain radiographs. On F-PET scans, 2 patients showed a diminished area with low uptake, 4 were unchanged, and 5 had enlarged areas. INTERPRETATION: Bone metabolism surrounding a volume of bone with no metabolic activity changes dynamically even 5 years after surgery. The presence of bone with minor uptake of F-tracer, indicating low or no bone metabolism, with further progression in 5 of 11 cases leads us to conclude that resurfacing THA should be used restrictively.