High Pressure Quenched Glasses: unique structures and properties.

Zr-based metallic glasses are prepared by quenching supercooled liquid under pressure. These glasses are stable in ambient conditions after decompression. The High Pressure Quenched glasses have a distinct structure and properties. The pair distribution function shows redistribution of the Zr-Zr interatomic distances and their shift towards smaller values. These glasses exhibit higher density, hardness, elastic modulus, and yield stress. Upon heating at ambient pressure, they show volume expansion and distinct relaxation behavior, reaching an equilibrated state above the glass transition. These experimental results are consistent with an idea of pressure-induced low to high density liquid transition in the supercooled melt.

Cantharidins induce ER stress and a terminal unfolded protein response in OSCC.

Mortality and morbidity associated with oral squamous cell carcinoma (OSCC) remain unacceptably high with disfiguring treatment options and a death rate of 1 per hour in the United States. The approval of cituximab for advanced OSCC has been the only new treatment for these patients since the 1970s, although it has not significantly increased overall survival. To address the paucity of effective new therapies, we undertook a high-throughput screen to discover small molecules and natural products that could induce endoplasmic reticulum (ER) stress and enforce a terminal unfolded protein response (UPR) in OSCC. The terpenoid cantharidin (CNT), previously used to treat various malignancies in culture-specific medical practices for over 2,000 y, emerged as a hit. CNT and its analog, cantharidic acid, potently induced protein and gene expression profiles consistent with the activation of ER stress, the UPR, and apoptosis in OSCC cells. Murine embryonic fibroblasts null for the UPR-associated transcription factors Atf4 or Chop were significantly protected from CNT, implicating a key role for the UPR in the death response. These data validate that our high-throughput screen can identify novel modulators of UPR signaling and that such compounds might provide a new therapeutic approach to treating patients with OSCC.

Foreign antigenic peptides delivered to the tumor as targets of cytotoxic T cells.

Cytotoxic T cells (CTL) are readily activated by immunogenic peptides and they exert potent anti-tumor activity if the same peptides are displayed on class I major histocompatibility complex (MHC) of the tumor cells. A handful of tumor-associated antigens have been identified and many of them are weak antigens. As an alternative strategy, strongly antigenic foreign peptides are delivered to the tumor, marking them for CTL recognition. To establish the principle of this new strategy, in vitro and in vivo tumor destruction was tested with BALB/c CTL to L(d)-associated beta-galactosidase (beta-gal) peptide p876. In vitro, anti-p876 CTL destroyed tumor cells in a single-cell suspension or in 3-D tumor boluses when exogenous p876 was added. Exogenous IL-2 was required to sustain CTL activity for complete destruction of tumor boluses. In vivo, BALB/c mice were immunized with p876 and a CD4 activating Pan DR reactive epitope (PADRE). PADRE, which binds to several different MHC class II antigen and activates CD4 T cells, induced delayed-type hypersensitivity and stimulated T cell proliferation. Immunized mice were injected with tumor cells loaded with p876 and mixed with PADRE. Starting from the day after tumor injection, mice received five rounds of peptide injection at the tumor sites and all tumors were rejected. Injection with saline had no effect. Injection with PADRE had minor anti-tumor activity. Immunization and treatment with p876 alone was not protective. Therefore, by delivering CD4 and CD8 reactive foreign peptides to the tumor, peptide-specific T cells rejected the tumors as demonstrated by the in vitro and in vivo tests.

Prognostic factors in major salivary gland cancer.

OBJECTIVE: To identify features of major salivary gland cancers that are prognostic for disease-free survival. STUDY DESIGN: A retrospective study of 78 patients with major salivary gland cancer (64 parotid and 14 submandibular gland) who underwent surgery for definitive treatment from 1976 to 1996. A select group of patients also received adjuvant radiation (56%) and/or chemotherapy (13%). METHOD: Clinical and pathological risk factors were obtained from patients' charts and pathology reports. Age, gender, tumor site, T-stage, facial paralysis, histologic neck involvement, perineural invasion, and cancer grade were analyzed with respect to disease-free survival. The role of adjuvant treatment in terms of clinical outcome was also investigated. RESULTS: In our series, the 5-year disease-free survival was 65%. Examining clinical and histologic features one at a time, we found poorer prognosis was associated with submandibular tumors compared with parotid (P =.02), higher T-stage (P =.001), positive cervical nodes (P <.001), perineural invasion (P =.002), and high-grade or adenoid cystic tumors (P =.002). A multivariable analysis indicated that positive lymph nodes (P =.07) and perineural invasion (P =.03) were important histologic predictors of shorter disease-free survival. Receipt of both adjuvant radiation and cisplatin-based chemotherapy (P =.05) was an independent predictor of longer disease-free survival. CONCLUSION: Our study indicated that the presence of positive lymph nodes and perineural invasion is important independent predictors of disease-free survival. Our limited data also suggest that adjuvant chemotherapy and radiation therapy may improve disease-free survival.

Craniocervical necrotizing fasciitis: an 11-year experience.

OBJECTIVE: We review our experience and present our approach to treating craniocervical necrotizing fasciitis (CCNF). STUDY DESIGN: All cases of CCNF treated at Wayne State University/Detroit Receiving Hospital from January 1989 to April 2000 were reviewed. Patients were analyzed for source and extent of infection, microbiology, co-morbidities, antimicrobial therapy, hospital days, surgical interventions, complications, and outcomes. RESULTS: A review of 250 charts identified 10 cases that met the study criteria. Five cases (50%) had spread of infection into the thorax, with only 1 (10%) fatality. An average of 24 hospital days (7 to 45), 14 ICU days (6 to 21), and 3 surgical procedures (1 to 6) per patient was required. CONCLUSION: Aggressive wound care, broad-spectrum antibiotics, and multiple surgical interventions resulted in a 90% (9/10) overall survival and 80% (4/5) survival for those with thoracic extension. SIGNIFICANCE: This is the largest single institution report of CCNF with thoracic extension identified to date.

Modified clotting properties of fibrinogen in the presence of acetylsalicylic acid in a purified system.

To assess how treatment with acetylsalicylic acid (ASA) alters the fibrin network structure, clotting was initiated in purified fibrinogen incubated with ASA by adding thrombin. Clotting time and maximum absorbance of the fibrin aggregation curve were used to demonstrate the potential of fibrin generation. The results showed that the clotting properties of fibrinogen decreased and that the affinity of plasminogen to fibrin or thrombin inhibition by antithrombin increased if plasminogen or antithrombin, respectively, were present in the reaction system. The effect of ASA varied in a dose dependent manner. It was concluded that ASA may directly or indirectly confer positive or negative effects on the stability of the fibrin clot and that the balance between these effects may be regulated by the ASA dose.

Titanium mesh repair of the severely comminuted frontal sinus fracture.

BACKGROUND: Severely comminuted frontal sinus fractures are difficult to contour and immobilize. Frequently, plates or wires are inadequate in fixating all fragments together, resulting in less than optimal outcomes. Advancements in the development of biomaterials have now made titanium mesh a new option for the repair of severely comminuted fractures. METHODS: Fourteen patients with severely comminuted frontal sinus fractures were treated with titanium mesh from 1994 to 1999. The fractures were reduced and immobilized using a simple algorithm: (1) Isolated anterior table fractures were repaired with reduced bony fragments attached to titanium mesh. (2) Anterior table fractures with nasofrontal duct involvement were repaired by sinus obliteration and anterior wall reconstruction with reduced bony fragments attached to titanium mesh. (3) Anterior and posterior table fractures with cerebrospinal fluid leak or displacement were treated with the cranialization of the sinus and anterior wall reconstruction with reduced bony fragments attached to titanium mesh. RESULTS: Of the 14 patients treated, 12 were available for postoperative evaluation. Parameters such as nasal function, cranial nerve V and VII function, cosmesis, and complications (hardware extrusions, sinusitis, meningitis, osteomyelitis, mucopyocele, brain abscess, pneumocephalus, and cerebrospinal fluid leak) were evaluated. All patients had good function of the superior division of cranial nerves V and VII. Two patients (16%) had minor wound infections, which resolved under treatment with antibiotics. All had excellent cosmetic results as measured by postreduction radiographs and personal and family perceptions of forehead contour. CONCLUSION: Titanium mesh reconstruction of severely comminuted frontal sinus fractures has few complications while providing excellent forehead contour and cosmesis.

Counterion-dye staining method for DNA in agarose gels using crystal violet and methyl orange.

Sensitive and safe methods for visualization of DNA in agarose gels are described. 0.001% crystal violet dissolved in distilled water was used for DNA staining on agarose gels and it could detect as little as 16 ng of DNA (3 kb, pGem-7Zf/EcoRI) without destaining procedure. The detection limit is four times lower than that of ethidium bromide. To improve the sensitivity, we studied a counterion-dye staining method using methyl orange as a counterion-dye which contributes to reduce excessive background staining by crystal violet. Dye concentration, pH of staining solution, mixing molar ratio of two dyes, and staining times were optimized for the counterion-dye staining. By the staining with a mixed solution of 0.0025% crystal violet and 0.0005% methyl orange in distilled water, 8 ng of the 3 kb DNA in an agarose gel was detected within 30 min.

Phase I trial of intratumoral liposome E1A gene therapy in patients with recurrent breast and head and neck cancer.

PURPOSE: We conducted a Phase 1 study to determine the maximal tolerated dose and maximum biologically active dose of the E1A gene delivered by intratumoral injection as a lipid complex with 3 beta[N-(n',n'-dimethylaminoethane)-carbamoyl] cholesterol/dioleoylphosphatidyl-ethanolamine (tgDCC-E1A). The E1A adenovirus gene functions as a tumor inhibitor gene by repressing oncogene transcription; modulating gene expression, resulting in cellular differentiation; and inducing apoptosis of cancer cells. E1A also sensitizes cancer cells to chemotherapeutic drugs such as etoposide, cisplatin, and taxol. EXPERIMENTAL DESIGN: Nine patients with recurrent and unresectable breast cancer and nine patients with head and neck cancer were enrolled. One tumor nodule in each patient was injected with tgDCC-E1A. Safety, tumor response, E1A gene transfer, and down-regulation of HER-2/neu were evaluated. RESULTS: No dose-limiting toxicity was observed in the four dose groups (15, 30, 60, and 120 microg DNA/cm of tumor). All patients tolerated the injections, although several experienced pain and bleeding at the injection site. A maximally tolerated dose was not reached in this study. E1A gene transfer was demonstrated in 14 of 15 tumor samples tested, and down-regulation of HER-2/neu was demonstrated in two of the five patients who overexpressed HER-2/neu at baseline. HER-2/neu could not be assessed in other posttreatment tumor samples because of extensive necrosis. In one breast cancer patient, no pathological evidence of tumor was found on biopsy of the treated tumor site at week 12. In 16 patients evaluable for tumor response, 2 had minor responses, 8 had stable disease, and 6 had progressive disease. CONCLUSIONS: Gene therapy with an E1A gene:lipid complex appears to be safe and warrants further testing.

Constructing deservingness: federal welfare reform, supplemental security income, and elderly immigrants.

On August 22, 1996, President Clinton signed the welfare reform law that ended eligibility for all immigrants to federal means tested entitlements. Poor elderly immigrants on Supplemental Security Income were specifically targeted. This article documents how the print media responded to these policy changes. The following are the major research questions: (1) How were older immigrants on Supplemental Security Income portrayed in the print media before and after federal welfare reform? (2) Who was involved in the print media coverage of older immigrants on Supplemental Security Income before and after federal welfare reform? (3) What types of statements were made by those involved in the print media coverage of older immigrants on Supplemental Security Income, before and after federal welfare reform? The approach used was an in-depth content analysis of newspaper articles from major U.S. newspapers. The findings demonstrate that older immigrants were constructed as "undeserving" in news articles prior to the passage of the federal welfare reform bill. However, after the passage of the federal welfare reform bill the coverage of older immigrants on Supplemental Security Income started to change, and older immigrants were portrayed as "deserving." In advancing aging policy for poor, vulnerable elderly, such as elderly immigrants, advocates, health and social service providers can play an influential role in bringing their voices to the print media.

Locally advanced nasopharyngeal cancer.

The Head and Neck Cancer Intergroup phase III clinical trial (Int 0099) for patients with locally advanced, squamous cell carcinomas (SCC) of the nasopharynx (or NPC) has been recently completed in the United States. The results of this study have defined the new standard of treatment for the group of patients studied. Patients with untreated, locally advanced stages III and IV NPC were randomized to a conventional course of radiation, or to radiation given concurrently with chemotherapy followed by three courses of combination chemotherapy. The 3-year progression-free survival (PFS) and overall survival (OS) were 24% versus 69% (P < 0.001) and 46% versus 76% (P < 0.001) for the control and experimental groups, respectively. Recent updates of these survival figures show that they have not changed appreciably. The considerable improvement in OS versus PFS for the patient group receiving radiation alone is accounted for primarily by re-treatment with concurrent radiation-chemotherapy, combination chemotherapy, and isolated salvage neck dissections. Highly significant differences in local control (41% vs 14%) and distant metastases (35% vs 13%) were demonstrated in favor of the chemoradiation treatment arm. The median age for these patients was 51 years, with a 2:1 male to female ratio. Although many patients had a significant history of tobacco exposure with or without alcohol use or abuse, only 24% had keratinizing or well-differentiated squamous (World Health Organization I) type tumors. Whether these results can be extrapolated to the more common Asian variety (WHO II and III) of advanced NPC must be addressed in future clinical trials.

Progression of head and neck cancer in an in vitro model.

OBJECTIVE: To identify alterations in angiogenesis and cell cycle regulation as preneoplastic cells progress to cancer in an in vitro model of head and neck tumor progression. METHODS: Immortal human gingival keratinocyte (IHGK) cells (preneoplastic) were derived from normal oral keratinocytes and were immortalized with human papillomavirus 16. Transformation of IHGK cells with a carcinogen (NNK, 4-[methylnitrosamino]-1-[3-pyridyl]-1-butanone) gave rise to IHGKN cells. We determined the growth rates, cell cycle phase, expression of cell cycle regulators, and expression of vascular endothelial growth factor along with the organotypic features of these cells and compared them with characteristics of head and neck cancer cells. RESULTS: IHGK and IHGKN cells grown in raft culture were morphologically similar to severe dysplasia and carcinoma, respectively. The proportion of cells in G(0)/G(1) was similar between IHGK and IHGKN. However, the proportion of IHGK cells was 35% greater in S phase as compared with the IHGKN cells, while a greater percentage (40%) of IHGKN cells were in G(2)/M. The expression of the other cell cycle regulators tested was unchanged. IHGK cells secreted less vascular endothelial growth factor on day 1 when compared with IHGKN (50.6 vs 245.6 pg/mL), along with a lower overall production rate (79% vs 133%). CONCLUSIONS: Transformation of IHGK cells resulted in the activation of vascular endothelial growth factor associated with angiogenesis. Inactivation of the G(1) cell cycle regulation occurred during immortalization and before transformation, and was sustained after carcinogen exposure. These alterations correspond to changes observed in patients with head and neck squamous cell carcinoma. This model can be useful in testing novel therapeutic and preventive strategies.

Neutron radiation enhances cisplatin cytotoxicity independently of apoptosis in human head and neck carcinoma cells.

Recent advances in combined modality treatment of locally advanced head and neck cancer have improved local and regional disease control and survival with better functional outcome. However, the local and regional failure rate after radiation therapy is still high for tumors that respond poorly to cisplatin-based neoadjuvant chemotherapy. This clinical observation suggests a common biological mechanism for resistance to cisplatin and photon irradiation. In this report, we investigated the molecular basis underlying cisplatin resistance in head and neck squamous carcinoma (HNSCC) cells and asked if fast neutron radiation enhances cisplatin cytotoxicity in cisplatin-resistant cells. We found that cisplatin sensitivity correlates with caspase induction, a cysteine proteinase family known to initiate the apoptotic cell death pathway, suggesting that apoptosis may be a critical determinant for cisplatin cytotoxicity. Neutron radiation effectively enhanced cisplatin cytotoxicity in HNSCCs including cisplatin-resistant cells, whereas photon radiation had little effect on cisplatin cytotoxicity. Interestingly, neutron-enhanced cisplatin cytotoxicity was associated neither with apoptosis nor with cell cycle regulation, as determined by caspase activity assay, annexin V staining, and flow cytometric analysis. Taken together, the present study provides a molecular insight into cisplatin resistance and may also provide a basis for more effective multimodality protocols involving neutron radiation for patients with locally advanced head and neck cancer.

Anti-allergic and anti-inflammatory triterpenes from the herb of Prunella vulgaris.

The activity-guided fractionation of the extract of the herb of Prunella vulgaris (Labiatae) led to the isolation of four triterpenes, i.e., betulinic acid, ursolic acid, 2 alpha,3 alpha-dihydroxyurs-12-en-28-oic acid, and 2 alpha-hydroxyursolic acid. One of these compounds, 2 alpha,3 alpha-dihydroxyursolic acid, demonstrated significant inhibition on the release of beta-hexosaminidase from the cultured RBL-2H3 cells in a dose-dependent manner; the IC50 value was calculated to be 57 microM. When the isolated compounds were tested for their effects on the production of nitric oxide from cultured murine macrophages, RAW 264.7 cells, ursolic acid and 2 alpha-hydroxyursolic acid exhibited strong inhibitory activities (IC50 values, 17 and 27 microM, respectively).

Anti-CD3/anti-CD28 bead stimulation overcomes CD3 unresponsiveness in patients with head and neck squamous cell carcinoma.

OBJECTIVES: To test whether T-cell CD3 responses are altered in patients with advanced-stage head and neck squamous cell carcinoma (HNSCC) and whether anti-CD3/anti-CD28 (alphaCD3/alphaCD28) bead stimulation could reverse CD3 unresponsiveness. DESIGN: Anti-CD3 (alphaCD3) monoclonal antibody immobilized on tissue culture plastic was used to stimulate lymph node mononuclear cells (LNMCs) and peripheral blood mononuclear cells (PBMCs) from patients with advanced-stage HNSCC. Proliferation, T-cell phenotype, and cytokines were measured during 8-day in vitro stimulation. Immune-enhancing properties of alphaCD3/ alphaCD28 beads were also tested on LNMCs and PBMCs. Cytotoxicity of bead-activated T cells (ATCs) was measured against autologous and allogeneic HNSCC. RESULTS: Six patients were nonresponders to alphaCD3 stimulation defined by tritium (3H) incorporation of less than 3500 cpm, whereas 11 patients were responders with 3H incorporation of 3500 cpm or more. Responders produced higher levels of interleukin (IL)-12 and interferon gamma (IFN-gamma) after alphaCD3 stimulation than nonresponders. No phenotypic or clinical differences were identified between groups. Stimulation with alphaCD3/alphaCD28 beads enhanced IFN-gamma and IL-2 produced by both groups. Bead ATCs were generated from PBMCs of patient 11 in the responder group and lysed (+/- SD) 100% +/-1% of autologous tumor and 49% +/-1% of allogeneic tumor. Bead ATCs from LNMCs of this patient lysed 58%+/-1% of autologous tumor and 63%+/-1% of allogeneic tumor. CONCLUSIONS: A subpopulation of patients with HNSCC who are nonresponders to alphaCD3 stimulation has been identified, showing reduced proliferation and IL-12 and IFN-gamma secretion. Nonresponders stimulated with alphaCD3/alphaCD28 beads reversed immune unresponsiveness and induced a type 1 cytokine response. Bead-generated ATCs from patient 11 in the responder group lysed autologous and allogeneic HNSCC in vitro, suggesting a possible effective immunotherapeutic modality in the treatment of HNSCC.

Direct Blue 71 staining of proteins bound to blotting membranes.

A sensitive staining method for protein blots using Direct Blue 71 is described. It is based on the selective binding of dye molecules to proteins in acidic solution and produces bluish violet colored bands. It is a simple and rapid procedure, involving only staining and rinsing steps that occur within 7 min. The sensitivity of this method is 5-10 ng of protein on nitrocellulose (NC) and 10-20 ng on polyvinylidene difluoride (PVDF), which is tenfold better than that of the commonly used Ponceau S staining. Moreover, the staining is reversible for subsequent immunostaining, without impairing immunoreactivity. To remove the dye from the developed bands, changes in pH and hydrophobicity of the solvent are required. Due to its sensitivity, rapidity, simplicity, and low cost, this stain may be more practical than other dye-based stains or metal-based stains for routine laboratory purposes.

Assessment of carotid artery invasion in patients with head and neck cancer.

PURPOSE: Define radiological and histological features in which patients with head and neck cancer would benefit from a carotid artery resection. Resection of the carotid artery has been advocated for local control of advanced squamous cell carcinoma of the head and neck. To provide appropriate preoperative counseling and optimize the utilization of resources, the criteria for patient selection has to be defined. METHODS: Thirty-four patients underwent carotid artery resection based on the clinical impression of tumor fixation. Eighteen and 28 patients were evaluated using computed tomography (CT) and histological analysis, respectively. The distance between the tumor cells and external elastic lamina was measured. CT scans were examined to determine the circumference of tumor attachment around the carotid artery. RESULTS: Clinical assessment predicted tumor within 1.8 mm of the carotid artery in 68% of cases. The overall survival for patients with tumor greater than 1.8 mm (N = 9) was better than that of patients with less (N = 19) than 1.8 mm (33.3% vs. 5.3%; median 24 versus 9 mo, P = .0899). Three of six patients (50%) with less than 180 degrees circumference tumor attachment had tumor within 1.8 mm from the external elastic lamina. Eight of twelve patients (67%) with tumors encompassing more than 180 degrees of the artery wall had tumor within 1.8 mm from the external elastic lamina. The overall survival rates for patients with tumor attachment greater and less than 180 degrees were 8.3% and 33%, respectively. DISCUSSION: Tumor invasion into the carotid artery was the strongest predictor of outcome. Clinical assessment was as predictive as CT for tumor invasion. If tumor involvement of the carotid artery is less than 180 degrees, peeling the tumor is an alternative to carotid artery resection.

Gene promoter hypermethylation in tumors and serum of head and neck cancer patients.

Promoter hypermethylation is an important pathway for repression of gene transcription in cancer cells. We analyzed aberrant DNA methylation at four genes in primary tumors from 95 head and neck cancer patients and then used the presence of this methylation as a marker for cancer cell detection in serum DNA. These four genes were tested by methylation-specific PCR and included: p16 (CDKN2A), O6-methylguanine-DNA-methyltransferase, glutathione S-transferase P1, and death-associated protein kinase (DAP-kinase). Fifty-five % (52 of 95) of the primary tumors displayed promoter hypermethylation in at least one of the genes studied: 27% (26/95) at p16, 33% (31 of 95) at O6-methylguanine-DNA-methyltransferase; and 18% (17 of 92) at DAP-kinase. No promoter hypermethylation was observed at the glutathione S-transferase P1 gene promoter. We detected a statistically significant correlation between the presence of DAP-kinase gene promoter hypermethylation and lymph node involvement (P = 0.014) and advanced disease stage (P = 0.016). In 50 patients with paired serum available for epigenetic analysis, the same methylation pattern was detected in the corresponding serum DNA of 21 (42%) cases. Among the patients with methylated serum DNA, 5 developed distant metastasis compared with the occurrence of metastasis in only 1 patient negative for serum promoter hypermethylation (P = 0.056). Promoter hypermethylation of key genes in critical pathways is common in head and neck cancer and represents a promising serum marker for monitoring affected patients.