Delayed diagnosis of oral lymphoma: a case series.

Lymphomas are the second most common neoplasm in the head and neck. The clinical and radiographic presentation of non-Hodgkin lymphoma in the oral cavity is non-specific and can be hard to differentiate from other common infectious or inflammatory conditions. We report four cases of lymphoma of the head and neck, which presented as odontogenic infection, osteomyelitis, or cutaneous infection, and subsequently led to a delay in provision of appropriate treatment. Correlation between clinical, radiographic and histological findings is essential in reaching an accurate diagnosis. It is important for clinicians to consider malignant lesions, such as lymphoma, in the differential diagnosis of pain, swelling, tooth mobility or radiographic radiolucencies. Clinicians should maintain a high level of suspicion for malignancy when inflammatory lesions fail to respond to normal modes of treatment.

Integration of high-throughput analytics and cell imaging enables direct early productivity and product quality assessment during Chinese Hamster ovary cell line development for a complex multi-subunit vaccine antigen.

Mammalian cell line generation typically includes stable pool generation, single cell cloning and several rounds of clone selection based on cell growth, productivity and product quality criteria. Individual clone expansion and phenotype-based ranking is performed initially for hundreds or thousands of mini-scale cultures, representing the major operational challenge during cell line development. Automated cell culture and analytics systems have been developed to enable high complexity clone selection workflows; while ensuring traceability, safety, and quality of cell lines intended for biopharmaceutical applications. Here we show that comprehensive and quantitative assessment of cell growth, productivity, and product quality attributes are feasible at the 200-1,200 cell colony stage, within 14 days of the single cell cloning in static 96-well plate culture. The early cell line characterization performed prior to the clone expansion in suspension culture can be used for a single-step, direct selection of high quality clones. Such clones were comparable, both in terms of productivity and critical quality attributes (CQAs), to the top-ranked clones identified using an established iterative clone screening approach. Using a complex, multi-subunit antigen as a model protein, we observed stable CQA profiles independently of the cell culture format during the clonal expansion as well as in the batch and fed-batch processes. In conclusion, we propose an accelerated clone selection approach that can be readily incorporated into various cell line development workstreams, leading to significant reduction of the project timelines and resource requirements.

Sarcoma of the proximal portion of the tibia in a dog 5.5 years after tibial plateau leveling osteotomy.

Osseous neoplasia was identified in the proximal portion of the tibia and distal portion of the femur in an 11.75-year-old spayed female German Shepherd Dog. A tibial plateau leveling osteotomy, followed by application of a metal plate, had been performed on the affected limb 5.5 years earlier. Areas of osteolysis and periosteal proliferation were seen radiographically, with an intense area of osteolysis directly beneath the metal plate. Histologically, the tumor was identified as a poorly differentiated sarcoma. Extracellular and intracellular debris was seen histologically, and energy-dispersive x-ray analysis confirmed that this debris was metallic. On visual examination, areas of the underside of the metal plate that had been in contact with bone had a dull, roughened appearance, and scanning electron microscopy of these areas revealed multiple corrosion pits. The plate was strongly magnetic, suggesting that it contained ferrite, and metallographic examination of the plate revealed substantial differences in the chemical makeup of various parts of the plate. Microstructure analysis revealed that the plate consisted of an austenite matrix with a large fraction of ferrite. The plate was determined to be a cast 316L stainless steel implant, but it did not meet American Society for Testing Materials standards for implant-grade materials. The possibility that implant corrosion might have played a role in tumor development is of concern; however, a definitive association was not established.

Neutrophil defense in patients undergoing bone marrow transplantation: bactericidal/permeability-increasing protein (BPI) and defensins in graft-derived neutrophils.

BACKGROUND: Even after neutrophil counts return to near normal levels, patients undergoing myeloablative chemotherapy and bone marrow transplantation (BMT) are at risk for invasive bacterial infections, raising the possibility that their neutrophil function might be impaired. To assess potential qualitative defects in neutrophil function in patients undergoing BMT, we measured neutrophil content of the antimicrobial (poly)peptides BPI and defensins. METHODS: Neutrophil extracts were analyzed for content of BPI by Western blotting and ELISA and for defensin peptides by acid-urea polyacrylamide gel electrophoresis (PAGE). Antibacterial activity of neutrophil extracts was measured against Escherichia coli K1/r, a clinical isolate sensitive to synergistic killing by BPI and defensins. RESULTS: Neutrophil extract BPI content on post-BMT days +20, +30, and +100 (169+/-35, 232+/-57, and 160+/-55 ng per 106 neutrophils, respectively) was similar to the neutrophil BPI content of normal controls (163+/-35 ng per 106 neutrophils). Neutrophil defensin content also did not vary during this time-span. Activity of neutrophil extracts against E. coli K1/r did not differ between BMT patients and controls. CONCLUSION: At post-BMT days +20 to +100, neutrophils derived from engrafted marrow contain normal quantities of BPI and defensins. Any deficiencies of neutrophil function during this phase are not due to inadequate expression of these antimicrobial (poly)peptides but could reflect abnormalities in other aspects of neutrophil function.

Evasion from NK cell immunity by MHC class I chain-related molecules expressing colon adenocarcinoma.

Evasion of host immune responses is well documented for viruses and may also occur during tumor immunosurveillance. The mechanisms involve alterations in MHC class I expression, Ag processing and presentation, chemokine and cytokine production, and lymphocyte receptor expression. Epithelial tumors overexpress MHC class I chain-related (MIC) molecules, which are ligands for the activating receptor NKG2D on NK and T cells. We report that NK cells from patients with colorectal cancer lack expression of activating NKG2D and chemokine CXCR1 receptors, both of which are internalized. Serum levels of soluble MIC (sMIC) are elevated and are responsible for down-modulation of NKG2D and CXCR1. In contrast, high serum levels of CXC ligands, IL-8, and epithelial-neutrophil-activating peptide (ENA-78) do not down-modulate CXCR1. In vitro, internalization of NKG2D and CXCR1 occurs within 4 and 24 h, respectively, of incubating normal NK cells with sMIC-containing serum. Furthermore, natural cytotoxicity receptor NKp44 and chemokine receptor CCR7 are also down-modulated in IL-2-activated NK cells cocultured in MIC-containing serum-an effect secondary to the down-modulation of NKG2D and not directly caused by physical association with sMIC. The patients' NK cells up-regulate expression of NKG2D, NKp44, CXCR1, and CCR7 when cultured in normal serum or anti-MIC Ab-treated autologous serum. NKG2D(+) but not NKG2D(-) NK cells are tumoricidal in vitro, and in vivo they selectively traffic to the xenografted carcinoma, form immunological synapse with tumor cells, and significantly retard tumor growth in the SCID mice. These results suggest that circulating sMIC in the cancer patients deactivates NK immunity by down-modulating important activating and chemokine receptors.

Lipid mediator-induced expression of bactericidal/ permeability-increasing protein (BPI) in human mucosal epithelia.

Epithelial cells which line mucosal surfaces are the first line of defense against bacterial invasion and infection. Recent studies have also indicated that epithelial cells contribute significantly to the orchestration of ongoing inflammatory processes. Here, we demonstrate that human epithelial cells express bactericidal/permeability-increasing protein (BPI), an antibacterial and endotoxin-neutralizing molecule previously associated with neutrophils. Moreover, we demonstrate that such BPI expression is transcriptionally regulated by analogs of endogenously occurring anti-inflammatory eicosanoids (aspirin-triggered lipoxins, ATLa). Initial studies to verify microarray analysis revealed that epithelial cells of wide origin (oral, pulmonary, and gastrointestinal mucosa) express BPI and each is similarly regulated by aspirin-triggered lipoxins. Studies aimed at localization of BPI revealed that such expression occurs on the cell surface of cultured epithelial cell lines and dominantly localizes to epithelia in human mucosal tissue. Functional studies employing a BPI-neutralizing anti-serum revealed that surface BPI blocks endotoxin-mediated signaling in epithelia and kills Salmonella typhimurium. These studies identify a previously unappreciated "molecular shield" for protection of mucosal surfaces against Gram-negative bacteria and their endotoxin.

Critical role of the complement system in group B streptococcus-induced tumor necrosis factor alpha release.

Group B Streptococcus (GBS) is a major cause of newborn sepsis and meningitis and induces systemic release of tumor necrosis factor alpha (TNF-alpha), believed to play a role in morbidity and mortality. While previous studies have shown that GBS can induce TNF-alpha release from monocytes and macrophages, little is known about the potential modulating effect of plasma or serum on GBS-induced TNF-alpha release, and there are conflicting reports as to the host receptors involved. In a human whole-blood assay system, GBS type III COH-1 potently induced substantial monocyte TNF-alpha release in adult peripheral blood and, due to a higher concentration of monocytes, 10-fold-greater TNF-alpha release in newborn cord blood. Remarkably, GBS-induced TNF-alpha release from human monocytes was enhanced approximately 1000-fold by heat-labile serum components. Experiments employing C2-, C3-, or C7-depleted serum demonstrated that C3 activation via the alternative pathway is crucial for potent GBS-induced TNF-alpha release. Accordingly, whole blood from C3-deficient mice demonstrated significantly reduced GBS-induced TNF-alpha release. Preincubation with human serum enhanced the TNF-alpha-inducing activity of GBS in a C3- and factor B-dependent manner, implying deposition of complement components via the alternative pathway. GBS-induced TNF-alpha release was inhibited by monoclonal antibodies directed against each of the components of CR3 and CR4: the common integrin beta subunit CD18 and the alpha subunits CD11b (of CR3) and CD11c (of CR4). Blood derived from CR3 (CD11b/CD18)-deficient mice demonstrated a markedly diminished TNF-alpha response to GBS. We conclude that the ability of plasma and serum to greatly amplify GBS-induced TNF-alpha release reflects the activity of the alternative complement pathway that deposits fragments on GBS and thereby enhances CR3- and CR4-mediated monocyte activation.