Jimmy J. Brown

A subpopulation of CD133+ cancer stem-like cells characterized in human oral squamous cell carcinoma confer resistance to chemotherapy

The potential role of cancer stem-like cells (CSCs) in chemoresistance of human oral squamous cell carcinoma (OSCC) was examined. A small subpopulation (1-2%) of CD133(+) CSCs was identified in OSCC cell lines and tissues. These CD133(+) CSCs possess higher clonogenicity, invasiveness, and increased in vivo tumorigenicity as compared to CD133(-) counterparts. Meanwhile, CD133(+) CSCs were substantially resistant to standard chemotherapy, wherein both in vitro and in vivo treatment with paclitaxel resulted in a marked enrichment for CD133(+) CSCs. Our data suggest that CD133(+) cells represent a small subpopulation of CSCs that may contribute to chemoresistance in human OSCC.

Resveratrol inhibits hypoxia-induced accumulation of hypoxia-inducible factor-1α and VEGF expression in human tongue squamous cell carcinoma and hepatoma cells

Hypoxia-inducible factor-1α (HIF-1α) is overexpressed in many human tumors and their metastases, and is closely associated with a more aggressive tumor phenotype. In this study, we investigated the effect of resveratrol, a natural product commonly found in grapes and various other fruits, on hypoxia-induced HIF-1α protein accumulation and vascular endothelial growth factor (VEGF) expression in human tongue squamous cell carcinomas and hepatoma cells. Our results showed that resveratrol significantly inhibited both basal level and hypoxia-induced HIF-1α protein accumulation in cancer cells, but did not affect HIF-1α mRNA levels. Pretreatment of cells with resveratrol significantly reduced hypoxia-induced VEGF promoter activities and VEGF expression at both mRNA and protein levels. The mechanism of resveratrol inhibition of hypoxia-induced HIF-1α accumulation seems to involve a gradually shortened half-life of HIF-1α protein caused by an enhanced protein degradation through the 26S proteasome system. In addition, resveratrol remarkably inhibited hypoxia-mediated activation of extracellular signal-regulated kinase 1/2 and Akt, leading to a marked decrease in hypoxia-induced HIF-1α protein accumulation and VEGF transcriptional activation. Functionally, we observed that resveratrol also significantly inhibited the hypoxia-stimulated invasiveness of cancer cells. These data suggested that HIF-1α/VEGF could be a promising drug target for resveratrol in the development of an effective chemopreventive and anticancer therapy in human cancers.

Overexpression of Human Papillomavirus Type 16 Oncoproteins Enhances Hypoxia-Inducible Factor 1α Protein Accumulation and Vascular Endothelial Growth Factor Expression in Human Cervical Carcinoma Cells

Purpose: Human papillomavirus (HPV)-16 oncoproteins, E6 and E7, are associated with enhanced tumor angiogenesis in human cervical cancers. The purpose of this study was (a) to investigate whether expression of HPV-16 E6 and E7 oncoproteins induces hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor expression in cervical cancer cells; and (b) to assess the effect of resveratrol on 16 E6- and E7-induced HIF-1α and VEGF gene expression. Experimental Design: Human cervical cancer cell lines C-33A and HeLa were transiently cotransfected with pSG5-HPV-16 E6 or 16 E7 constructs along with HIF-1α small interfering RNA (siRNA) or nonspecific siRNA. The expression of HIF-1α/VEGF was measured using real-time PCR, Western blot analysis, or ELISA. The in vitro angiogenic activity induced by 16 E6- and E7-transfected cells was examined. The effect of resveratrol on oncoprotein-induced HIF-1α/VEGF expression and in vitro angiogenesis was investigated. Results: HPV-16 E6- and E7-transfected cervical cancer cells express increased HIF-1α protein and VEGF expression. These stimulatory effects were abrogated by cotransfection with either HIF-1α siRNA or treatment with resveratrol. Blocking extracellular signal-regulated kinase 1/2 (ERK 1/2) and phosphoinositide-3-kinase by PD98059 and LY294002, respectively, abolished 16 E6- and E7-induced HIF-1α and VEGF expression. Functionally, we showed that HPV-16 E6- and E7-transfected cervical cancer cells stimulated in vitro capillary or tubule formation, and these angiogenic effects could be abolished either by cotransfection with HIF-1α siRNA or by treatment with resveratrol. Conclusion: HPV-16 oncoproteins contribute to enhanced angiogenesis in cervical cancer cells via HIF-1α–dependent VEGF expression. Resveratrol suppresses 16 E6- and E7-induced HIF-1α–mediated angiogenic activity and, thus, is a promising chemotherapeutic agent for human cervical cancer.

Treatment with siRNA and antisense oligonucleotides targeted to HIF-1α induced apoptosis in human tongue squamous cell carcinomas

Overexpression of hypoxia inducible factor‐1α (HIF‐1α) in cancers has been correlated to a more aggressive tumor phenotype. We investigated the effect of HIF‐1α knockout on the in vitro survival and death of human tongue squamous cell carcinomas (SCC‐4 and SCC‐9). Under normoxic condition, a basal level of HIF‐1α protein was constitutively expressed in SCC‐9 cells, albeit an undetectable level of HIF‐1α messages. Exposure to hypoxia induced only a transient increase in mRNA transcript but a prolonged elevation of HIF‐1α protein and its immediate downstream target gene product, VEGF. Under normoxic or hypoxic conditions, treatment of SCC‐9 cells with AS‐HIF‐1α ODN suppressed both constitutive and hypoxia‐induced HIF‐1α expression at both mRNA and protein levels. Knockout of HIF‐1α gene expression via either AS‐HIF‐1α ODN or siRNA (siRNAHIF‐1α) treatment resulted in inhibition of cell proliferation and induced apoptosis in SCC‐4 and SCC‐9 cells. We also demonstrated that exposure of SCC‐9 cells to hypoxia led to a time‐dependent increase in the expression of bcl‐2 and IAP‐2, but not p53. The attenuated levels of bcl‐2 and IAP‐2, and the enhanced activity of caspase‐3 after treatment with AS‐HIF‐1α ODN may contribute partly to the effects of HIF‐1α blockade on SCC‐9 cell death. Collectively, our data suggest that a constitutive or hypoxia‐induced expression of HIF‐1α in SCC‐9 and SCC‐4 cells is sufficient to confer target genes expression essential for tumor proliferation and survival. As a result, interfering with HIF‐1α pathways by antisense or siRNA strategy may provide a therapeutic target for human tongue squamous cell carcinomas.

Otolaryngologic Manifestations of Down Syndrome

The DS patient has clear anatomic differences in the head and neck region when compared with the general population. These anomalies include a flat occiput, oblique palpebral fissures, epicanthal folds, speckled irides, a protruding tongue, prominent malformed ears, and a flat nasal bridge. Congenital otologic anomalies and acquired conditions such as otitis media are also more frequently observed in the DS population. The DS patient is predisposed to obstructive sleep apnea, and the diagnosis of sleep apnea in DS patients is more likely to be delayed. A child with DS who has a narrowed nasopharynx, large tongue, and a subglottis which is smaller than normal must be given special consideration at the time of intubation. Such a patient requires an endotracheal tube two sizes smaller than the standard size appropriate for the patients age. The child should also be suspected of having and be evaluated for obstructive sleep apnea, to ensure that appropriate precautions are taken in the perioperative period. Finally, any DS patient undergoing preoperative evaluation for a general anesthetic should have a careful assessment of the cervical spine to avoid dislocation or spinal cord injury. Hearing loss may be suspected in any congenital syndrome. In DS, there is a clearly increased incidence of congenital temporal bone anomalies, external auditory canal stenosis, and otitis media. All DS patients should undergo hearing assessment in the neonatal period, with follow-up as appropriate. Aggressive treatment of conductive hearing loss and early amplification may be necessary to maximize speech and language development.

Tumor-Like Stem Cells Derived from Human Keloid Are Governed by the Inflammatory Niche Driven by IL-17/IL-6 Axis

Background Alterations in the stem cell niche are likely to contribute to tumorigenesis; however, the concept of niche promoted benign tumor growth remains to be explored. Here we use keloid, an exuberant fibroproliferative dermal growth unique to human skin, as a model to characterize benign tumor-like stem cells and delineate the role of their “pathological” niche in the development of the benign tumor. Methods and Findings Subclonal assay, flow cytometric and multipotent differentiation analyses demonstrate that keloid contains a new population of stem cells, named keloid derived precursor cells (KPCs), which exhibit clonogenicity, self-renewal, distinct embryonic and mesenchymal stem cell surface markers, and multipotent differentiation. KPCs display elevated telomerase activity and an inherently upregulated proliferation capability as compared to their peripheral normal skin counterparts. A robust elevation of IL-6 and IL-17 expression in keloid is confirmed by cytokine array, western blot and ELISA analyses. The altered biological functions are tightly regulated by the inflammatory niche mediated by an autocrine/paracrine cytokine IL-17/IL-6 axis. Utilizing KPCs transplanted subcutaneously in immunocompromised mice we generate for the first time a human keloid-like tumor model that is driven by the in vivo inflammatory niche and allows testing of the anti-tumor therapeutic effect of antibodies targeting distinct niche components, specifically IL-6 and IL-17. Conclusions/Significance These findings support our hypothesis that the altered niche in keloids, predominantly inflammatory, contributes to the acquirement of a benign tumor-like stem cell phenotype of KPCs characterized by the uncontrolled self-renewal and increased proliferation, supporting the rationale for in vivo modification of the “pathological” stem cell niche as a novel therapy for keloid and other mesenchymal benign tumors.

Nondestructive Imaging of Live Human Keloid and Facial Tissue Using Multiphoton Microscopy

OBJECTIVES To use multiphoton microscopy to image collagen fibers and matrix structure in nonfixed human keloid tissue and normal human facial skin obtained following surgery and to compare the findings to existing knowledge of normal skin and keloid morphology to determine if this technology is a suitable adjunct for conventional histology. METHODS Epidermis was removed to expose the fibroblast-rich dermal layer that was then imaged using a multiphoton confocal microscope (Zeiss-Meta 510; Carl Zeiss, Jena, Germany). An 800-nm tunable titanium/sapphire femtosecond laser (Mai-Tai; Newport Co Spectra-Physics, Mountain View, California) was used to excite the tissue; second harmonic generation between 397 and 408 nm and autofluorescent signals were collected. Images were obtained using a Plan-Neofluar x40 oil immersion objective lens and a Plan-Apochromat x63 oil immersion lens. RESULTS Compared with normal skin, keloids showed disorganized collagen fibers arranged in complex swirls and bundles 20 to 30 microm in diameter. Normal tissue showed collagen fibers as distinct, straight strands less than 10 microm in diameter. Differences between normal and keloid tissue were subtle but apparent. CONCLUSIONS The value of imaging living tissue is a significant benefit. Because keloids and hypertrophic scars result from altered collagen metabolism, the development of clinical multiphoton microscopy systems may allow examination of wound healing dynamics in vivo and potentially provides a means to monitor therapy without the need for biopsy or the risk of injury to tissue.

Carcinoma of the oral pharynx: an analysis of subsite treatment heterogeneity

The data indicate that SCC of the various subsites of the oropharynx can be treated successfully with acceptable locoregional control and survival rates by using either surgery or primary radiotherapy for TI or T2 primary lesions. Treatment success data for late-stage disease (T3 and T4) are less encouraging. regardless of which modality is used or which treatment center is administering treatment. This finding may suggest an intrinsic property of these lesions or the patient that may be going unnoticed.One problem is that the diversity of approaches to these lesions hinders any meaningful comparisons between series from different treatment centers. There exists heterogeneity in patient populations and approaches to staging and characterization of these diseases. This situation has ensured the same heterogeneity in treatment philosophy, which is largely institutionally based.

Phagocytosis of candida albicans by lymphatic tumour cells in vitro

Experiments were carried out to investigate whether different lymphatic tumour cell lines have similar kinetic characteristics of phagocytosis of microorganisms. Six tumour cell lines were used. These were a human T-cell line (CEM), a mouse T-cell line (YAC-1), a human B-cell line (LAZ), and a human erythroleukemic tumour cells (K562), whereas 2 cell lines of professional phagocytosis were used as controls, a human macrophage cell line (THP1) and a mouse macrophage cell line (P388D1). Tumour cells were mixed with candida albicans at a ratio of 10:1 of candida to tumour cells and the percentage of tumour cells that had attached/phagocytosed candida was determined. After 4 h coculture with candida, tumour cells not of T-cell origin (LAZ and K562) showed moderate level of phagocytosis (28%), whereas tumour cells of T-cell origin (CEM and YAC-1) demonstrated low levels of phagocytosis (15%) as compared to macrophage cell lines (THP1 and P388D1) that showed maximum phagocytosis (64-78%). Acid phosphatase (AcPase) activity was increased by 33% during coculture of YAC-1 cells and yeast cells. In conclusion, the results suggest that lymphatic tumour cells of nonphagocytic origin acquire phagocytic properties during the course of malignancy, and digestion of phagocytosed yeast cells maybe related with AcPase activity, as well as that of other lysosomal enzymes. This phenomenon may represent one mechanism by which tumour cells downregulate immune surveillance.

Landmarks for endoscopic approach to the parapharyngeal internal carotid artery: A radiographic and cadaveric study

To define transnasal endoscopic surgical landmarks for the parapharyngeal segment of the internal carotid artery (ppICA) using radiographic analysis and cadaveric dissection.