Craniomaxillofacial osteosarcoma - The role of surgical margins.

BACKGROUND: Osteosarcomas (OS) in the craniomaxillofacial (CMF) region are typically diagnosed at later age than long-bone OS, but they are reported to have better 5-year survival. Curative treatment warrants wide surgical resection, which is often not possible in the CMF region. The purpose of this article is to present a nationwide series of CMF in Finland to discuss the role of surgery. PATIENTS AND METHODS: All 21 CMF OS patients managed in Finland from 1992 to 2009 were included. The mean age was 40 years (range 15-72). Data on patient and tumor characteristics, treatment modalities, and survival were recorded. All patients had a minimum follow-up of 5 years or until death. RESULTS: OS was evenly represented in the mandible and maxillary bones, which together constituted 76% of all sites. Surgery with curative intent was carried out in 20 patients. Clear margins were achieved in only five cases. Eight (40%) of these 20 patients died due to OS, and their average survival time was 1.3 years. Seven (35%) out of the 20 patients received radiotherapy due to close/intralesional surgical margins, and four of them did not develop recurrences during the follow-up. CONCLUSIONS: The results suggest that postoperative radiotherapy may alter the prognosis in CMF OS, particularly in cases with close or intralesional margins. This may increase the survival rates achieved by prompt action in performing radical surgery.

Role of two sequence motifs of mesencephalic astrocyte-derived neurotrophic factor in its survival-promoting activity.

Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a prosurvival protein that protects the cells when applied intracellularly in vitro or extracellularly in vivo. Its protective mechanisms are poorly known. Here we studied the role of two short sequence motifs within the carboxy-(C) terminal domain of MANF in its neuroprotective activity: the CKGC sequence (a CXXC motif) that could be involved in redox reactions, and the C-terminal RTDL sequence, an endoplasmic reticulum (ER) retention signal. We mutated these motifs and analyzed the antiapoptotic effect and intracellular localization of these mutants of MANF when overexpressed in cultured sympathetic or sensory neurons. As an in vivo model for studying the effect of these mutants after their extracellular application, we used the rat model of cerebral ischemia. Even though we found no evidence for oxidoreductase activity of MANF, the mutation of CXXC motif completely abolished its protective effect, showing that this motif is crucial for both MANF's intracellular and extracellular activity. The RTDL motif was not needed for the neuroprotective activity of MANF after its extracellular application in the stroke model in vivo. However, in vitro the deletion of RTDL motif inactivated MANF in the sympathetic neurons where the mutant protein localized to Golgi, but not in the sensory neurons where the mutant localized to the ER, showing that intracellular MANF protects these peripheral neurons in vitro only when localized to the ER.

Primary metastatic Ewing's family tumors: results of the Italian Sarcoma Group and Scandinavian Sarcoma Group ISG/SSG IV Study including myeloablative chemotherapy and total-lung irradiation.

BACKGROUND: The Italian Sarcoma Group and the Scandinavian Sarcoma Group designed a joint study to improve the prognosis for patients with Ewing's family tumors and synchronous metastatic disease limited to the lungs, or the pleura, or a single bone. PATIENTS AND METHODS: The study was opened in 1999 and closed to the enrollment in 2008. The program consisted of intensive five-drug combination chemotherapy, surgery and/or radiotherapy as local treatment, and consolidation treatment with high-dose busulfan/melphalan plus autologous stem cell rescue and total-lung irradiation. RESULTS: During the study period, 102 consecutive patients were enrolled. The median follow-up was 62 months (range 24-124). The 5-year event-free survival probability was 0.43 [standard deviation (SD) = 0.05] and the 5-year overall survival probability was 0.52 (SD = 0.052). Unfavorable prognostic factors emerging on multivariate analysis were a poor histological/radiological response at the site of the primary tumor [relative risk (RR) = 3.4], and incomplete radiological remission of lung metastases after primary chemotherapy (RR = 2.6). One toxic death and one secondary leukemia were recorded. CONCLUSIONS: This intensive approach is feasible and long-term survival is achievable in ∼50% of patients. New treatment approaches are warranted for patients responding poorly to primary chemotherapy.

Determination of residual antiplatelet activity of clopidogrel before neuraxial injections.

BACKGROUND: Guidelines recommend discontinuation of clopidogrel for 7 days before a neuraxial injection, while other directives suggest that 5 days might be adequate. We examined the time course of antiplatelet activity after clopidogrel discontinuation in patients undergoing epidural injections. METHODS: Thirteen patients were studied at baseline, 3, 5, and 7 days after discontinuation of clopidogrel. P(2)Y(12) determinations were performed using the VerifyNow(®) assay (Accumetrics, San Diego, CA, USA), and clot closure times with stimulation by collagen/epinephrine and collagen/adenosine diphosphate using the PFA-100(®) (Platelet Function Analyzer, Siemens Diagnostics, Deerfield, IL, USA). Repeated-measures ANOVA was used to evaluate P(2)Y(12) platelet reaction units, PFA-100 closure times, and per cent P(2)Y(12) inhibition values. Wilcoxon's signed-rank test was used to compare the frequencies of ≥30%, 11-29%, and ≤10% platelet inhibition between the baseline and subsequent sampling points after discontinuation of clopidogrel. RESULTS: On day 3 after clopidogrel discontinuation, two subjects had ≥30%, seven subjects had 11-29%, and four subjects had ≤10% platelet inhibition; the corresponding numbers were 0, 3, and 10 subjects on day 5 (P=0.04). There were no differences between the ≥30%, 11-29%, and <10% platelet inhibition groups between days 5 and 7 (0, 0, and 13 subjects, P=1.0). PFA-ADP closure times were normal throughout the study period except in one patient. CONCLUSIONS: These findings support the recommendation that discontinuation of clopidogrel for 5 days allows >70% of platelet function and might be adequate before a neuraxial injection is performed.

Expression of GATA-6 transcription factor in pleural malignant mesothelioma and metastatic pulmonary adenocarcinoma.

BACKGROUND: Malignant mesothelioma (MM) is a highly aggressive tumour with poor prognosis and limited response to therapy. New markers for the prediction of prognosis in MM and in pulmonary adenocarcinoma of the pleura are valuable. GATA-6 belongs to a six member zinc finger transcription factor family named after their recognition motif W-GATA-R. AIM: To clarify the distribution and possible function of GATA-6 transcription factor in MM and in pleural metastasis of lung adenocarcinomas. METHODS: 63 pleural MM and 36 pleural metastatic pulmonary adenocarcinomas were studied for GATA-6 expression by immunohistochemistry using tissue microarrays. Expression of GATA-6 was examined in relation to thyroid transcription factor-1 expression, survival, proliferation and apoptosis. RESULTS: Nuclear immunoreactivity for GATA-6 was stronger and more frequent in MM than in metastatic pleural adenocarcinoma. Prognosis was better in patients with GATA-6 expression when compared to those with no GATA-6 expression (p = 0.002); in the subgroup analysis the difference was significant in epithelial and sarcomatous mesothelioma. GATA-6 was not associated with spontaneous proliferation or apoptosis of the tumour cells in situ. CONCLUSION: Results suggest that GATA-6 plays a role in pleural malignancies, predicting longer survival in subgroups of MM.

Gene copy number analysis in malignant pleural mesothelioma using oligonucleotide array CGH.

Conventional cytogenetic analyses and comparative genomic hybridization have revealed a complex and even chaotic nature of chromosomal aberrations in pleural malignant mesothelioma (MM). We set out to describe the complex gene copy number changes and screen for novel genetic aberrations using a high-density oligonucleotide microarray platform for comparative genomic hybridization (aCGH) of a series of 26 well-characterized MM tumor samples. The number of copy number changes varied from zero to 40 per sample. Gene copy number losses predominated over gains, and the most frequent region of loss was 9p21.3 (17/26 cases), the locus of CDKN2A and CDKN2B, both known to be commonly lost in MM. The most recurrent minimal regions of losses were 1p31.1--> p13.2, 3p22.1-->p14.2, 6q22.1, 9p21.3, 13cen-->q14.12, 14q22.1-->qter, and 22qcen-->q12.3. Previously unreported gains included 9p13.3, 7p22.3-->p22.2, 12q13.3, and 17q21.32-->qter. The results suggest that gene copy number losses are a major mechanism of MM carcinogenesis and reveal a recurrent pattern of copy number changes in MM.

Etiology of specific molecular alterations in human malignancies.

Cancer results from multiple genomic changes that affect DNA and its gene expression. The DNA sequences may be gained, lost or amplified, or translocated into different parts of the genome to form a fusion gene with oncogenic properties. The occurrence of specific chromosomal aberrations may be restricted to only one cancer type and it may be considered a primary carcinogenic event. Furthermore, the aberration profiles may be used to cluster tumors with similar origins. A variety of techniques exist for the detection of specific chromosomal and gene expression changes. However, the etiology of these molecular alterations remains unclear. Here we discuss the roles of Helicobacter pylori and asbestos burden as carcinogens that cause gastric cancer, mesothelioma and lung cancer.

Alveolar gas composition before and after maximal breath-holds in competitive divers.

The urge to breathe, as stimulated by hypercapnia, is generally considered to cause a breath-hold diver to end the breath-hold, and pre-breath hold hyperventilation has been suggested to cause hypoxic loss of consciousness (LOC) due to the reduced urge to breathe. Competitors hyperventilate before "Static Apnea", yet only 10% surface with symptoms of hypoxia such as loss of motor control (LMC) or LOC. We hypothesized that the extensive hyperventilation would prevent hypercapnia even during prolonged breath-holding and we also recorded breaking-point end-tidal PO2 in humans. Nine breath-hold divers performed breath-holds of maximal duration according to their chosen "Static Apnea" procedure. They floated face down in a swimming pool (28 degrees C). The only non-standard procedure was that they exhaled into a sampling tube for end-expiratory air, before starting the breath-hold and before resuming breathing. Breath-hold duration was 284 +/- 25 (SD) seconds. End-tidal PCO2 was 18.9 +/- 2.0 mmHg before apnea and 38.3 +/- 4.7 mmHg at apnea termination. End-tidal PO2 was 131.7 +/- 2.7 mmHg before apnea and 26.9 +/- 7.5 mmHg at apnea termination. Two of the subjects showed LMC after exhaling into the sampling tube; their end-tidal PAO2 values were 19.6 and 21.0 mmHg, respectively. End-tidal CO2 was normocapnic or hypocapnic at the termination of breath-holds. These data suggest that the athletes rely primarily on the hypoxic stimuli, probably in interaction with CO2 stimuli to determine when to end breath-holds. The severity of hypoxia close to LOC was similar to that reported for acute hypobaric hypoxia in humans.

Effects of IKK inhibitor PS1145 on NF-kappaB function, proliferation, apoptosis and invasion activity in prostate carcinoma cells.

A key antiapoptotic transcription factor, nuclear factor kappa-B (NF-kappaB), is known to be critically important for tumor cell growth, angiogenesis and development of metastatic lesions. We and others showed previously that NF-kappaB transcription factor was constitutively activated in androgen-independent prostate carcinoma (PC) cell lines due to the upregulated activity of inhibitor of NF-kappaB kinases (IKK). In this work, using luciferase assay, electrophoretic mobility shift assay and Northern blot analysis of expression of endogenous kappaB-responsive genes, we demonstrate that a novel highly specific small-molecule IKK inhibitor, PS1145, efficiently inhibited both basal and induced NF-kappaB activity in PC cells. We found that PS1145 induced caspase 3/7-dependent apoptosis in PC cells and significantly sensitized PC cells to apoptosis induced by tumor necrosis factor alpha. We also showed that PS1145 inhibited PC cell proliferation. Effects of PS1145 on proliferation and apoptosis correlated with inhibition of interleukin (IL)-6, cyclin D1, D2, inhibitor of apoptosis (IAP)-1 and IAP-2 gene expression and decreased IL-6 protein level. In addition, we found that incubation with PS1145 inhibited the invasion activity of highly invasive PC3-S cells in invasion chamber assay in a dose-dependent manner. Overall, this study provides the framework for development of a novel therapeutic approach targeting NF-kappaB transcription factor to treat advanced PC.

Lactation and breast-feeding ability following lateral pedicle mammaplasty.

A retrospective study was undertaken to evaluate the ability of breast-feeding using lateral pedicle reduction mammaplasty. A standard questionnaire was sent to 72 patients who had been operated on at a fertile age. Data was collected regarding the duration and quality of preoperative and postoperative breast-feeding, difficulties while breast-feeding, reasons for discontinuation of breast-feeding or for not attempting to breast-feed, and postoperative subjective sensitivity. Thirteen of the women who replied to the questionnaire had given birth after surgery. The pressure threshold sensitivity of areola-nipple complex was measured in nine of the above cases. To preserve lactation, a technique leaving structures untouched within the pedicle with increased dimensions was used.Breast-feeding was considered successful in this study if it was performed exclusively, without supplementation, for two months. Seven women (54%) breast-fed successfully for between two and 14 months (5.8+/-1.3) following surgery. Two women (16%) were classified as unsuccessful and four women (30%) did not breast-feed at all. The success of breast-feeding was limited by nonsurgical factors including the influence of medical personal. Five of eight women (62%) who had children preoperatively improved their rate of breast-feeding after surgery. No correlation was demonstrated between measured sensitivity and breast-feeding (p=0.65). No significant correlation was found between the resected tissue and breast-feeding (p=0.08). No relation was observed between the duration of breast-feeding and the period between operation and partus.

A retrospective quantitative assessment of breast sensation after lateral pedicle mammaplasty.

Preservation of sensitivity is one of the important objectives in reduction mammaplasty. The lateral or medial pedicle reduction mammaplasty technique aims to maintain superficial innervation of the breast. These superficial nerves are smaller and more variable compared with deep and larger nerve structures, which are excised during this procedure. An assessment of recovery of tactile sensitivity after lateral pedicle mammaplasty was performed retrospectively. One hundred and one women, operated on between the year 1985 and 2000, with a lateral pedicle mammaplasty, were reviewed. Using a standardised questionnaire, women reported on subjective changes in sensation after surgery, including time of recovery and the influence which surgery had on sensual function. Fifty-nine of these women were tested 3-7 years after surgery. Touch sensitivity was measured using a set of 20 Semmes-Weinstein monofilaments. Erectile function was also tested. The mean pressure threshold for the areola region was 19.12 g/mm(2) and for the nipple 16.75 g/mm(2), which is better that in studies on inferior pedicle mammaplasty. In previous studies patients were tested earlier, which may explain this difference.No correlation was found between sensitivity and resected tissue (for the areola p=0.88 and for the nipple p=0.82) or between sensitivity and age of patients (for the areola p=0.73 and for the nipple p=0.80). There were individual differences in the results. It is speculated that variability in anatomy of superficial nerves might explain these differences. Nearly all patients tested regained their erectile function. The majority of patients did not notice any change in their erogenous function. Twelve (12%) reported a post-surgical improvement of sensation. Nipples with partial loss of sensation persisted in 2% of the patients.

Small, novel proteins from the mistletoe Phoradendron tomentosum exhibit highly selective cytotoxicity to human breast cancer cells.

Four novel proteins (phoratoxins C-F) have been isolated from the North American mistletoe Phoradendron tomentosum. The amino acid sequences of these phoratoxins were determined unambiguously using a combination of Edman degradation and trypsin enzymatic digestion, and by electrospray ionization tandem mass spectrometry sequencing. Phoratoxins C, E and F consist of 46 amino acid residues; and phoratoxin D of 41. All proteins had six cysteines, similar to the earlier described phoratoxins A and B, which are thionins. The cytotoxicity of each protein was evaluated in a human cell line panel that represented several cytotoxic drug-resistance mechanisms. For the half-maximal inhibitory concentrations (IC50 values) of the different cell lines in the panel, correlation with those of standard drugs was low. The most potent cytotoxic phoratoxin C was further tested on primary cultures of human tumor cells from patients. The solid tumor samples from breast cancer cells were 18 times more sensitive to phoratoxin C than the tested hematological tumor samples.

A novel multiparametric approach for analysis of cytoplasmic immunoglobulin light chains by flow cytometry.

We describe a novel flow cytometric approach using a two-step acquisition technique to determine the cytoplasmic immunoglobulin light chains (LC) expression. Samples were prepared by a lysed-whole-blood technique and incubated with CD38-PE (phycoerythrin) and CD45-FITC (fluorescein isothiocyanate). The cells were fixed and acquired on an FACSCalibur flow cytometer (first acquisition). The cells were then permeabilized, incubated with either kappa-FITC or lambda-FITC and reacquired (second acquisition). Analysis of the data was performed by gating on the differing intensities of CD38 and evaluating them for the presence of a shifting FITC-positive population from the first acquisition to the second acquisition. The FITC fluorescence intensity of the second acquisition was equal to the sum of surface CD45 expression obtained during the first acquisition and the cytoplasmic LC expression obtained during the second acquisition. Thus, the shifting (increase) of FITC fluorescence intensity during the second acquisition is specifically due to the cytoplasmic expression of either the kappa or lambda LC. We studied 15 multiple myeloma (MM) patients and 10 controls (samples from patients without plasma cell dyscrasias). None of the controls showed evidence of any clonal populations. Thirteen of 15 MM patients exhibited clonal plasma cells (CD38 bright), ranging from 0.01% to 34% of total events collected. In addition, we identified another minute clonal population of lymphocytes (CD38 dim, CD45 bright, low forward and side scatter) in 12 of 13 MM patients with clonal plasma cells. This population, ranging from 0.01% to 0.6% of total events collected, had the same LC restriction as the clonal plasma cells. Patients with a ratio of minor clonal population to clonal plasma cells less than 0.07 tended to remain in partial or complete remission than those with a ratio > or =0.07 (4/5 versus 1/4, P <.1, chi(2)). We conclude that this method is highly sensitive and permits us to identify the minute clonal population of lymphocytes in MM patients. Our preliminary observations with a small cohort of patients imply that this minute clonal population may have important prognostic significance. The prognostic significance should be confirmed by further studies.

Cytotoxicity of digitoxin and related cardiac glycosides in human tumor cells.

The saponin digitonin, the aglycone digitoxigenin and five cardiac glycosides were evaluated for cytotoxicity using primary cultures of tumor cells from patients and a human cell line panel (representing different cytotoxic drug-resistance patterns). Of these seven compounds, proscillaridin A was the most potent (IC(50): 6.4--76 nM), followed by digitoxin, and then ouabain, digoxin, lanatoside C, digitoxigenin and digitonin. Correlation analysis of the log IC(50) values for the cell lines in the panel showed that compound cytotoxicity was only slightly influenced by resistance mechanisms that involved P-glycoprotein, topoisomerase II, multidrug resistance-associated protein and glutathione-mediated drug resistance. Digitoxin and digoxin expressed selective toxicity against solid tumor cells from patients, while proscillaridin A expressed no selective toxicity against either solid or hematological tumor cells. The results revealed marked differences in cytotoxicity between the cardiac glycosides, both in potency and selectivity, and modes of action for cytotoxicity that differ from that of commonly used anticancer drugs.

Methods of cytologic smear preparation and fixation. Effect on the immunoreactivity of commonly used anticytokeratin antibody AE1/AE3.

OBJECTIVE: To evaluate the effect of fixation and methods of cytologic smear preparation on the immunoreactivity of commonly used anticytokeratin antibody AE1/AE3. STUDY DESIGN: Scrape cytology smears and formalin-fixed, paraffin-embedded tissue sections (FPTS) of 20 unfixed, fresh specimens submitted for intraoperative consultation were studied by the immunoperoxidase method. In addition to the morphologic examination, the smears and FPTS were evaluated for intensity and proportion scores. For each specimen, two scrape cytology smears were wet fixed in 95% ethanol, and 12 smears were air dried without fixation. Air-dried smears were either postfixed after rehydration in saline or fixed directly without rehydration by one of the three fixatives: alcoholic formalin, 95% ethanol with 5% acetic acid or 95% ethanol. RESULTS: Both intensity and proportion scores were higher with rehydrated, air-dried smears as compared to those without rehydration and were comparable to those with wet-fixed smears and FPTS. In the rehydrated group, the optimum results were achieved when the smears were postfixed with alcoholic formalin. CONCLUSION: The method of preparation and fixation had variable effects on the immunoreactivity of anticytokeratin antibody AE1/AE3. The optimum results were achieved with saline-rehydrated, air-dried smears post-fixed in alcoholic formalin. To evaluate the role of inter-sample variation, further, larger studies are recommended on this and other antibodies before applying them to different types of cytologic smears.

A novel human processed gene, DAD-R, maps to 12p12 and is expressed in several organs.

A cDNA of a processed gene of human DAD-1 (defender against apoptotic cell death) was cloned from the human neuroblastoma cell line SH-SY5Y. The genomic sequence of this novel processed gene, DAD-R, lacked introns and was flanked by 8 bp terminal repeats. RT-PCR showed that the transcript is expressed predominantly in testis, ovaries, pancreas, lung and skeletal muscle. DAD-R has several possible initiation codons, one of them producing an open reading frame comprising 75% of the DAD-1 gene. We determined the chromosomal localization of DAD-R as 12p11.2-12p12.1, an area linked to familial synpolydactyly and frequently amplified in a variety of cancers, including those of testis, ovaries, pancreas and lungs.

Experience in qualitative and quantitative FDG PET in follow-up of patients with suspected recurrence from head and neck cancer.

We evaluated positron emission tomography (PET) with 2-[fluorine-18]fluoro-2-deoxy-D-glucose (FDG) in the detection of recurrent head and neck cancer, and compared visual and quantitative interpretation of PET images for their accuracy in the identification of tumour recurrence. Sixty-two FDG PET studies were performed in 56 patients having a total of 81 lesions, which were clinically suspected for recurrent carcinoma of the head and neck. The PET images were interpreted visually, and tracer uptake was quantitated as the standardised uptake value adjusted to body weight (SUV). Sensitivity of visual interpretation of the PET images for the presence of malignancy ranged from 84 to 95%, and specificity from 84 to 93%, respectively, depending on the selected scheme for grading of the lesions. Malignant lesions accumulated significantly more FDG than the benign ones (the median SUVs were 6.8 and 3.3, respectively, P<0.001). However, there was a wide overlap of the FDG uptake values between these two groups. Hence, the highest accuracy of quantitative analysis in correct identification of tumour recurrence (75% at Receiver Operating Curve analysis) was inferior to that of visual analysis (89%). FDG PET is feasible for the detection of recurrent head and neck cancer. Although quantitation of FDG uptake using SUV shows significantly higher tracer concentrations for malignant than benign lesions, the wide overlap of individual SUVs between these two groups is a serious concern in diagnostic evaluation. Therefore, in clinical practice it may be preferable to identify the presence of tumour recurrence within this patient group by qualitative interpretation of the PET images.

Glycosylation of phytepsin and expression of dad1, dad2 and ost1 during onset of cell death in germinating barley scutella.

Dysfunction and downregulation of dad (defending against death) has been linked to programmed cell death (PCD) in animals and plants. As DAD is an essential subunit of the oligosaccharyltransferase that is located in the ER membrane, the results have raised the possibility that downregulation of N-linked glycosylation could be involved in the regulation of PCD. Here we show that the 16 kDa subunit of phytepsin, a vacuolar proteinase, is normally processed and glycosylated at the onset of DNA fragmentation in germinating barley scutella. Two cDNA clones encoding dad (dad1, dad2), and one cDNA encoding another subunit of the same oligosaccharyltransferase complex (ost1) were isolated from barley. Northern analysis of germinating scutella show that the expression of only dad1 is declining before onset of DNA fragmentation. In contrast to this, the expression of both dad2 and ost1 increase before onset of DNA fragmentation.