Different cellular p16(INK4a) localisation may signal different survival outcomes in head and neck cancer.

BACKGROUND: Recently, the management of head and neck squamous cell carcinoma (HNSCC) has focused considerable attention on biomarkers, which may influence outcomes. Tests for human papilloma infection, including direct assessment of the virus as well as an associated tumour suppressor gene p16, are considered reproducible. Tumours from familial melanoma syndromes have suggested that nuclear localisation of p16 might have a further role in risk stratification. We hypothesised p16 staining that considered nuclear localisation might be informative for predicting outcomes in a broader set of HNSCC tumours not limited to the oropharynx, human papilloma virus (HPV) status or by smoking status. METHODS: Patients treated for HNSCC from 2002 to 2006 at UNC (University of North Carolina at Chapel Hill) hospitals that had banked tissue available were eligible for this study. Tissue microarrays (TMA) were generated in triplicate. Immunohistochemical (IHC) staining for p16 was performed and scored separately for nuclear and cytoplasmic staining. Human papilloma virus staining was also carried out using monoclonal antibody E6H4. p16 expression, HPV status and other clinical features were correlated with progression-free (PFS) and overall survival (OS). RESULTS: A total of 135 patients had sufficient sample for this analysis. Median age at diagnosis was 57 years (range 20-82), with 68.9% males, 8.9% never smokers and 32.6% never drinkers. Three-year OS rate and PFS rate was 63.0% and 54.1%, respectively. Based on the p16 staining score, patients were divided into three groups: high nuclear, high cytoplasmic staining group (HN), low nuclear, low cytoplasmic staining group (LS) and high cytoplasmic, low nuclear staining group (HC). The HN and the LS groups had significantly better OS than the HC group with hazard ratios of 0.10 and 0.37, respectively, after controlling for other factors, including HPV status. These two groups also had significantly better PFS than the HC staining group. This finding was consistent for sites outside the oropharynx and did not require adjustment for smoking status. CONCLUSION: Different p16 protein localisation suggested different survival outcomes in a manner that does not require limiting the biomarker to the oropharynx and does not require assessment of smoking status.

Modulation of the metastatic phenotype by 13-cis-retinoic acid.

KLN 205 murine squamous carcinoma cells were grown in medium supplemented with the retinoid 13-cis-retinoic acid (RA) to study the relationship between RA-induced cell surface changes and alterations of the metastatic phenotype. Modulation of the cell surface glycoconjugate expression was measured by flow cytometric analysis of the RA-treated tumor cells stained with fluoresceinated lectins. RA treatment (5 X 10(-6) and 5 X 10(-7) M) altered the glycoconjugate expression of KLN 205 cells in a selective, dose-dependent fashion. Tumor cells grown in RA-supplemented medium for more than 4 days demonstrated greatly increased binding of fluoresceinated Griffonia simplicifolia I lectin, peanut lectin, wheat-germ lectin, concanavalin A, and soybean lectin (P less than .001), but the increased binding of Ulex europaeus lectin was of a much smaller magnitude (P = .02). After 15 days of growth in these noncytotoxic or cytostatic concentrations of RA, malignant KLN 205 cells had a greatly decreased proclivity to metastasize, as measured by the lung colony assay (P = .0003). The RA-induced cell surface glycoconjugate changes preceded the decrease in experimental metastatic potential. Since enzymatic (neuraminidase) alteration of the tumor cell surface to produce glycoconjugate expression similar to that seen in RA-treated cells also reduced the ability of the KLN 205 cells to form lung colonies (P = .0022), it is suggested that RA-induced alteration of the cell surface carbohydrate antigens is related to the decreased experimental metastatic potential seen in tumor cells treated with RA.

The effects of monocyte-conditioned medium and interleukin 1 on the synthesis of collagenous and non-collagenous proteins by mouse bone and human bone cells in vitro.

Cultured adherent human mononuclear cells produce factor(s) which stimulate the release of calcium from new-born mouse calvaria in organ culture. This stimulation of bone resorption is accompanied by an inhibition of the incorporation of [3H]proline into collagen which is independent of increased prostaglandin production by the bone. When human osteoblast-like cells are treated with conditioned medium from human mononuclear cells, collagen accounts for a decreased proportion of the protein synthesised. This effect on matrix synthesis is not accompanied by an inhibitory action of the monocyte-conditioned medium preparations on net cell proliferation. In human osteoblast-like cell cultures, partially purified human interleukin 1 also inhibits the production of the bone-specific protein osteocalcin in a dose-dependent fashion. These observations are consistent with the hypothesis that products of human monocytes similar to, or identical with, human interleukin 1 may be important regulators of bone metabolism and may contribute to the bone loss seen in diseases such as chronic rheumatoid arthritis.

Characterization of cells with high alkaline phosphatase activity derived from human bone and marrow: preliminary assessment of their osteogenicity.

Confluent cellular layers are reproducibly obtained (from 21 of 24 specimens) by outgrowth from composite pieces of human trabecular bone and marrow. The cells resemble fibroblasts in terms of morphology, esterase profile, and production of collagen type 1. However, the cells displayed some osteoblastlike features. Both the primary outgrowths and passaged cultures had high alkaline phosphatase activities (37 nmols min-1 X microgram DNA-1) in the range displayed by embryonic osteoblastlike cells. The cellular alkaline phosphatase activity, which showed similarity to the bone isoenzyme on kinetic criteria, was stimulated by 1,25-dihydroxyvitamin D3 but decreased by PTH (1-34). In addition, the cell preparations were shown to increase osteocalcin (bone Gla protein) production in response to 1,25-dihydroxyvitamin D3. The osteogenic potential of the bone and marrow-derived cells has been assessed in an in vivo diffusion chamber assay in which congenitally athymic (nude) mice were used as hosts. None of the 25 chambers examined showed evidence of osteogenesis, although the cells remained viable and fibroblastlike. The alkaline phosphatase activities decreased to less than 1% of the original, high in vitro values. The findings question the hypothesis that bone and marrow-derived cells are osteoblasts or osteoblastlike cells, rather than a mixture of cell lines of the bone and marrow stromal system.

Comparison of 19-lodocholesterol and 6-lodomethylnorcholesterol as adrenal-scanning agents.

The two adrenal-specific compounds, 19-iodocholest-5-en-3beta-ol (19-iodocholesterol) and 6beta-iodometyly-19-norcholest-5(10)-en-3beta-ol (6-iodomethylnorcholesterol), synthesized pure in gram amounts have been labeled with iodine-125 and used for tissue-distribution studies in the rat. Radioactivity from 6-iodomethylnorcholesterol is accumulated in the rat adrenal gland at least 18 times more than is radioactivity from 19-iodocholesterol at 24 hr, and 50 times more at 72 hr. The use of radioactive 6-iodomethylnorcholesterol gave superior adrenal images in rats at earlier times than did radioactive 19-iodocholesterol, and the former should be equally superior when used in humans.

Regional analysis of D2 dopamine receptors in Parkinson's disease using SPECT and iodine-123-iodobenzamide.

UNLABELLED: The goal of this study was to examine the relationship between D2 dopamine receptor density and levodopa dosage, disease duration and dyskinesia in Parkinson's disease (PD). METHODS: Iodine-123-iodobenzamide SPECT scans were obtained from 14 PD patients and 12 age-matched controls using a three-headed camera in conjunction with MRI and a fiducial-based image registration system to define regions of interest. Basal ganglia/cerebellum counts/voxel ratios in dorsal and ventral head of caudate and anterior and posterior putamen were measured at 30, 60, 120 and 180 min postinjection. As in 11C-raclopride studies, ratios obtained at that time when they asymptomatically approach a maximum value (180 min) were accepted as the best measure of receptor density. RESULTS: Among PD patients, a trend towards an inverse correlation between regional basal ganglia/cerebellum ratios and levodopa dosage achieved significance in ventral caudate (F = 6.244, p = 0.037); similarly, an inverse correlation between these ratios and disease duration achieved significance in anterior putamen (F = 13.144, p = 0.007). Ratios were significantly lower in anterior putamen in patients with dyskinesia (t = 3.068, p = 0.042). CONCLUSION: In PD, the previously observed inverse correlation between levodopa dosage and D2-receptor density appears to be most prominent in the least dopamine-depleted region, ventral caudate. There may be a genuine effect of disease duration on receptor density in putamen and reduced receptor density in anterior putamen may be associated with dyskinesia.

Biodistribution and radiation dosimetry of radioiodinated-SCH 23982, a potential dopamine D-1 receptor imaging agent.

Radioiodinated-SCH 23982 is a potential agent for the imaging of dopamine D-1 receptors in the human brain. In vivo binding of [125I]SCH 23982 to D-1 receptors in rat brain was determined over 4 hr. The ratio of activity in striatum and frontal cortex to that in cerebellum increased over the first 2 hr to maximum values of 4.4:1 and 2.1:1, respectively. The percent injected dose in whole brain at 0.5 and 2 hr were 0.62 and 0.15, respectively. Administration of the antagonists propranolol (beta-1), prazosin (alpha-1), haloperidol (D-2) and ketanserin (5HT-2) did not significantly alter the striatum/cerebellum ratio; however, SCH 23390, a D-1 antagonist, totally blocked ligand uptake by striatum and frontal cortex. Biologic distribution data in the rat were determined after injection of 3 microCi of [125I]SCH 23982. 76% of the injected dose was excreted in 48 hr via the liver and kidneys. Internal radiation absorbed dose estimates to nine source organs, total body, the GI tract, gonads and red bone marrow were calculated for humans using the physical decay data for 123I. The critical organ was found to be the lower large intestine which received 1.1 rad/mCi of the administered dose. The total-body dose was 63 mrad/mCi. The data indicate that [123I]SCH 23982 should be a suitable agent for imaging the D-1 dopamine receptor in the human brain by single photon emission computed tomography.

In vivo binding in rat brain and radiopharmaceutical preparation of radioiodinated HEAT, an alpha-1 adrenoceptor ligand.

In vivo binding of [125I]-2-[beta-(3-iodo-4-hydroxyphenyl)ethylaminomethyl tetralone) ([125I]HEAT) to alpha-1 adrenoceptors in the rat brain was determined over 4 hr. Uptake in the thalamus and frontal cortex was approximately 0.1% injected dose per gram tissue. Thalamus/cerebellum ratios of 10:1 and frontal cortex/cerebellum ratios of 5:1 were found at 4 hr. Pretreatment with prazosin, an alpha-1 antagonist, completely inhibited the accumulation of [125I]HEAT in thalamus and frontal cortex; yet uptake of radioactivity was not significantly affected by antagonists and agonists for other receptors classes (propranolol, beta-1; apomorphine, D-1; spiperone, D-2). Binding of [125I]HEAT is saturable. At 4 hr, [125I]HEAT or [123I]HEAT was shown to be the only radioactive material in rat thalamus and frontal cortex. Iodine-123 HEAT and [125I]HEAT were synthesized as radiopharmaceuticals within 3 hr in 99% radiochemical purity.

Radiation dosimetry of iodine-123 HEAT, an alpha-1 receptor imaging agent.

Biologic distribution data in the rat were obtained for the alpha-1 adrenoceptor imaging agent (+/-) 2-[beta-(iodo-4-hydroxyphenyl)ethylaminomethyl]tetralone (HEAT) labeled with [125I]. The major excretory routes were through the liver (67%) and the kidney (33%). Internal radiation absorbed dose estimates to nine source organs, total body, the GI tract, gonads, and red bone marrow were calculated for the human using the physical decay data for [123I]. The critical organ was found to be the lower large intestine, receiving 1.1 rad per mCi of [123I]HEAT administered. The total-body dose was found to be 58 mrad per mCi.

Vitamin D metabolites regulate osteocalcin synthesis and proliferation of human bone cells in vitro.

The effects of six natural vitamin D metabolites of potential biological and therapeutic interest, 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3), 25-hydroxyvitamin D3 (25-OH-D3), 24R,25-dihydroxyvitamin D3 (24R,25-(OH)2D3), 1,24R,25-trihydroxyvitamin D3 (1,24R,25-(OH)3D3), 25S,26-dihydroxyvitamin D3 (25S,26-(OH)2D3) and 1,25S,26-trihydroxyvitamin D3 (1,25S,26-(OH)3D3) on cell replication and expression of the osteoblastic phenotype in terms of osteocalcin production were examined in cultured human bone cells. At a dose of 5 X 10(-12) mol/1, 1,25-(OH)2D3 stimulated cell proliferation, whereas at higher doses (5 X 10(-9)-5 X 10(-6) mol/1) cell growth was inhibited in a dose-dependent manner. The same pattern of effects was seen for the other metabolites in a rank order of potency: 1,25-(OH)2D3 greater than 1,25S,26-(OH)3D3 = 1,24R,25-(OH)3D3 greater than 25S,26-(OH)2D3 = 24R,25-(OH)2D3 = 25-OH-D3. Synthesis of osteocalcin was induced by 1,25-(OH)2D3 in doses similar to those required to inhibit cell proliferation. Biphasic responses were observed for some of the metabolites in terms of osteocalcin synthesis, inhibitory effects becoming apparent at 5 X 10(-6) mol/1. The cells did not secrete osteocalcin spontaneously. These results indicate that vitamin D metabolites may regulate growth and expression of differentiated functions of normal human osteoblasts.

Risk factors for post-stem cell transplant sinusitis.

An understanding of the factors that place the post-transplant patient at increased risk for sinusitis would help identify patients likely to develop disease and possibly allow for interventions that would decrease the incidence or severity of sinus disease. This retrospective study investigates the ability of screening paranasal sinus computed tomographic scans (CTs), clinical history, and potential risk factors for sinusitis, including history of tobacco use, history of allergies or asthma, IgG level, history of sinusitis, remission status and acute graft-versus-host disease (GVHD) to predict post-transplant sinusitis. Medical records and sinus CTs of 100 allogeneic bone marrow recipients were reviewed. There was no increased risk of developing sinusitis post SCT for patients with significant disease on screening CT, symptoms at time of transplant, a history of tobacco use, asthma or allergies, low IgG level, history of sinusitis or for patients at high risk of relapse. Patients with GVHD were 4.3 times more likely than patients without GVHD to develop sinusitis post transplant (95% CI: 1.7-11.0, P = 0.002). Acute GVHD places patients at greater risk of developing sinus infections.

Excretion of octopamine metabolites in neuroblastoma.

The urinary concentrations of o-hydroxymandelic acid, m-hydroxymandelic acid, p-hydroxymandelic acid, homovanillic acid and vanillylmandelic acid were determined in 57 healthy children and 9 patients with neuroblastoma. The concentrations of o-hydroxymandelic acid and p-hydroxymandelic were not significantly different for both groups whereas the concentrations of m-hydroxymandelic acid, homovanillic acid and vanillylmandelic acid were elevated 20- to 30-fold in the neuroblastoma patients.

Changes in plasma osteocalcin concentration following treatment with stanozolol.

Treatment of healthy, adult, human volunteers with stanozolol, 5 mg twice daily, for six weeks caused a marked elevation in circulating levels of osteocalcin. Plasma osteocalcin concentrations did not fall to pretreatment values for a period in excess of four weeks after the cessation of treatment but remained similar to levels whilst on treatment. At the same time, as the rise in plasma osteocalcin was seen, plasma alkaline phosphatase activity fell; however, in this case, a return to pretreatment levels occurred within four weeks of treatment ending. These data demonstrate that stanozolol has a detectable effect on the plasma concentration of a protein thought to be a marker of osteoblast activity, and that this effect continues for a period after treatment has been discontinued.

Normal excretion of m-hydroxymandelic acid in hypertensive patients.

o-Hydroxymandelic acid (OHMA), m-hydroxymandelic acid (MHMA) and p-hydroxymandelic acid (PHMA) were measured in the urine of 42 normotensive and 54 hypertensive patients. Patients having high urinary MHMA levels were all found to be ingesting medications containing m-synephrine (phenylephrine). These patients also had high levels of urinary m-synephrine which was excreted as the glucuronide. When patients ingesting m-synephrine were excluded from the analysis, no significant differences were observed between the two groups for the urinary excretion of OHMA, MHMA and PHMA.

Synthesis and purification of radioactive 6beta-iodomethyl-19-norcholest-5(10)-en-3beta-ol.

A method for the synthesis and purification of 6beta-iodomethyl-19-norcholest-5(10)-ene-3beta-ol-131I of greater than 98 mole% chemical purity and greater than 99% radiochemical purity is presented. Carbon-13 and proton NMR were used to establish the identity and purity. Discrepancies in the characterization of this compound, previously published by two other research groups, are discussed.

Chemical and radiochemical stability of the adrenal-scanning agents, 6beta-iodomethyl-19-norcholest-5(10-en-3beta-ol and 19-iodocholest-5-en-3beta-ol.

The chemical stabilities of the adrenal-scanning agents, 6beta-iodo-methyl-19-norcholest-5(10)-en-3beta-ol (6-iodomethylnorcholesterol) and 19-iodocholest-5-en-3beta-ol (19-iodocholesterol), and several of their derivatives were examined by 13C nuclear magnetic resonance. Neat 6-iodomethylnorcholesterol, sealed in glass under nitrogen and stored at 0 degrees C, remains 98 mole% chemically pure for 3 months. Neat 19-iodocholesterol, stored in the dark at 25 degrees C, remains 98 mole% chemically pure for 3 months. Either 6-iodomethylnorcholesterol-125I or-131I, informulation and stored at 5 degrees C, will remain greater than 97% radiochemically pure for at least 15 days. Labelled 19-iodocholesterol, formulated and stored under the same conditions, shows 20% decomposition after 3 weeks and 40% after 6 weeks.

Efficacy of fiberoptic laryngoscopy in the diagnosis of inhalation injuries.

BACKGROUND: A significant proportion of burn patients with inhalation injuries incur difficulties with airway protection, dysphagia, and aspiration. In assessing the need for intubation in burn patients, the efficacy of fiberoptic laryngoscopy was compared with clinical findings and the findings of diagnostic tests, such as arterial blood gas analysis, measurement of carboxyhemoglobin levels, pulmonary function tests, and radiography of the lateral aspect of the neck. OBJECTIVE: To determine if these patients were at risk for aspiration or dysphagia, barium-enhanced fluoroscopic swallowing studies were performed. DESIGN: Prospective study. SETTINGS: Burn intensive care unit in an academic tertiary referral center. MAIN OUTCOME MEASURES: Need for endotracheal intubation and potential for aspiration. RESULTS: Six (55%) of 11 patients had clinical findings and symptoms that indicated, under traditional criteria, endotracheal intubation for airway protection. Visualization of the upper airway with fiberoptic laryngoscopy obviated the need for endotracheal intubation in all 11 patients. These patients also failed to evidence an increased risk of aspiration or other swallowing dysfunction. CONCLUSIONS: In comparison with other diagnostic criteria, fiberoptic laryngoscopy allows differentiation of those patients with inhalation injuries who, while at risk for upper airway obstruction, do not require intubation. These patients may be safely observed in a monitored setting with serial fiberoptic examinations, thus avoiding the possible complications associated with intubation of an airway with a compromised mucosalized surface. In these patients, swallowing abnormalities do not manifest.

Assessment of donor-site functional morbidity from radial forearm fasciocutaneous free flap harvest.

OBJECTIVE: To quantitate the functional morbidity to the hand and wrist following harvest of a radial forearm fasciocutaneous free flap. DESIGN: Prospective case-control study, with each patient providing his or her internal control, comparing preoperative and postoperative operated to nonoperated forearms. SETTING: Tertiary care hospital in large metropolitan area. PATIENTS: A consecutive sample of 11 patients who underwent a radial forearm free flap reconstruction of the head and neck from April 1997 to May 1998. MAIN OUTCOME MEASURES: Range of motion of the wrist (flexion and extension, ulnar and radial deviation), grip and pinch strength, and sharp and dull sensation in the distribution of the radial, ulnar, and median nerves. RESULTS: Statistically significant differences (P<.05) were measured in wrist flexion, pinch strength, and sharp sensation in the anatomical snuffbox of the operated forearm. No subjective complaints of loss of function were reported by any patient. CONCLUSIONS: Donor-site functional morbidity associated with harvest of the radial forearm fasciocutaneous free flap is measurable. The statistical differences found do not translate into subjective patient complaints of everyday functional morbidity.

Assessment of percutaneous fasciotomy in the management of Dupuytren's contracture.

This prospective study of the treatment of Dupuytren's Contracture in 78 hands has been designed to investigate the role of subcutaneous fasciotomy. The results suggest that in hands where the contracture is predominantly at the metacarpophalangeal joint, then percutaneous fasciotomy is of value.