Early endothelial damage and increased colonic vascular permeability in the development of experimental ulcerative colitis in rats and mice.

The role of endothelial damage and increased vascular permeability (VP) in the pathogenesis of ulcerative colitis (UC) has not been investigated. We examined using functional, morphologic, and molecular biologic studies whether and to what extent the endothelial barrier dysfunction precedes enhanced epithelial permeability (EP) and the development of mucosal lesions during the early stages of experimental UC. We showed that in rats with iodoacetamide (IA)-induced UC increased colonic VP occurs early (ie, 2.6-fold increase at 15 min, P<0.01) preceding changes in epithelial barrier permeability. EP was unchanged at 15 and 30 min after IA administration and was increased 1.9-fold at 1 h and 6.7-fold at 2 h (both P<0.001) after IA. In the dextran sodium sulfate-induced slowly developing UC, colonic VP was significantly increased in 2 days (P<0.05) and EP only in 4 days (P<0.05). Mucosal endothelial injury led to hypoxia (P<0.05) of colonic surface epithelial cells 30 min after IA administration that was associated with increased expressions of transcription factors hypoxia-inducible factor-1α and early growth response-1. Electron and light microscopy demonstrated areas of colonic mucosa with perivascular edema covered by intact layer of surface epithelial cells in both rat and mouse models of UC. This is the first demonstration in four models of UC that endothelial damage, increased colonic VP, perivascular edema, and epithelial hypoxia precede epithelial barrier dysfunction that is followed by erosions, ulceration, and inflammation in UC.

Lung cancer: a classic example of tumor escape and progression while providing opportunities for immunological intervention.

Lung cancers remain one of the most common and deadly cancers in the world today (12.5% of newly diagnosed cancers) despite current advances in chemo- and radiation therapies. Often, by the time these tumors are diagnosed, they have already metastasized. These tumors demonstrate the classic hallmarks of cancer in that they have advanced defensive strategies allowing them to escape various standard oncological treatments. Immunotherapy is making inroads towards effectively treating other fatal cancers, such as melanoma, glioblastoma multiforme, and castrate-resistant prostate cancers. This paper will cover the escape mechanisms of bronchogenic lung cancer that must be overcome before they can be successfully treated. We also review the history of immunotherapy directed towards lung cancers.

Changes in tumor-antigen expression profile as human small-cell lung cancers progress.

OBJECTIVE: Our group has previously observed that in patients with small-cell lung cancers (SCLCs), the expression of a tumor antigen, glioma big potassium (gBK) ion channel, is higher at the time of death than when the cancer is first treated by surgical resection. This study aimed to determine whether this dichotomy was common in other potential lung tumor antigens by examining the same patient samples using our more extensive profile analysis of tumor-antigen precursor protein (TAPP). We then tested the hypothesis that therapeutic intervention may inadvertently cause this increased gBK production. METHODS: SCLC samples (eight surgical resections and three autopsy samples) and three control lungs were examined by quantitative real-time polymerase chain reaction for 42 potential TAPPs that represent potential T-cell-mediated immunological targets. RESULTS: Twenty-two TAPP mRNAs displayed the same profile as gBK, i.e., more mRNAs were expressed at autopsy than in their surgical counterparts. B-cyclin and mouse double minute 2, human homolog of P53-binding protein were elevated in both autopsy and surgical specimens above the normal-lung controls. When HTB119 cells were incubated with doxorubicin, gBK was strongly induced, as confirmed by intracellular flow cytometry with a gBK-specific antibody. CONCLUSION: Our findings suggested that more immunological targets became available as the tumor responded to chemotherapy and proceeded toward its terminal stages.

HCA519/TPX2: a potential T-cell tumor-associated antigen for human hepatocellular carcinoma.

BACKGROUND: Immunotherapy for human hepatocellular cancer (HCC) is slowly making progress towards treating these fatal cancers. The identification of new antigens can improve this approach. We describe a possible new antigen, hepatocellular carcinoma-associated antigen-519/targeting protein for Xklp-2 (HCA519/TPX2), for HCC that might be beneficial for T-cell specific HCC immunotherapy. METHODS: HCC was studied for the expression for 15 tumor-associated antigens considered useful for immunotherapy within three HCC cell lines (HepG2, Hep3B, and PLC/PRF/5), lymphocytes, non-cancerous livers, and clinical HCC. The expression of tumor antigenic precursor proteins (TAPPs) messenger RNA was first screened by reverse transcriptase quantitative real-time polymerase chain reaction. RESULTS: Four antigens (alpha fetoprotein, aspartyl/asparaginyl ß-hydroxylase, glypican-3 and HCA519/TPX2) proved to be the best expressed TAPPs within the HCC specimens by molecular analyses. HCA519/TPX2 was detected by intracellular cell flow cytometry within HCC cell lines by using a specific antibody towards this TAPP. This antibody also detected the protein within primary HCCs. We synthesized two HCA519/TPX2 peptides (HCA519464-472 and HCA519351-359) which can bind to human leukocyte antigen (HLA)-A*0201. Dendritic cells pulsed with these peptides stimulated cytolytic T lymphocytes (CTLs). These killer T-cells lysed HLA-A*0201+ T2 cells exogenously loaded with the correct specific peptide. The CTLs killed HepG2 (HLA-A2+ and HCA519+), but not the Hep3B and PLC/PRF/5 cell lines, which are HCA519+ but HLA-A2-negative. In silico analysis reveals that HCA519/TPX2 has the inherent ability to bind to a very wide variety of HLA antigens. CONCLUSION: HCA519/TPX2 is a viable immunotarget that should be further investigated within HCC patients.

Small cell lung cancer cells express the late stage gBK tumor antigen: a possible immunotarget for the terminal disease.

Big Potassium (BK) ion channels have several splice variants. One splice variant initially described within human glioma cells is called the glioma BK channel (gBK). Using a gBK-specific antibody, we detected gBK within three human small cell lung cancer (SCLC) lines. Electrophysiology revealed that functional membrane channels were found on the SCLC cells. Prolonged exposure to BK channel activators caused the SCLC cells to swell within 20 minutes and resulted in their death within five hours. Transduction of BK-negative HEK cells with gBK produced functional gBK channels. Quantitative RT-PCR analysis using primers specific for gBK, but not with a lung-specific marker, Sox11, confirmed that advanced, late-stage human SCLC tissues strongly expressed gBK mRNA. Normal human lung tissue and early, lower stage SCLC resected tissues very weakly expressed this transcript. Immunofluorescence using the anti-gBK antibody confirmed that SCLC cells taken at the time of the autopsy intensely displayed this protein. gBK may represent a late-stage marker for SCLC. HLA-A*0201 restricted human CTL were generated in vitro using gBK peptide pulsed dendritic cells. The exposure of SCLC cells to interferon-γ (IFN-γ) increased the expression of HLA; these treated cells were killed by the CTL better than non-IFN-γ treated cells even though the IFN-γ treated SCLC cells displayed diminished gBK protein expression. Prolonged incubation with recombinant IFN-γ slowed the in vitro growth and prevented transmigration of the SCLC cells, suggesting IFN-γ might inhibit tumor growth in vivo. Immunotherapy targeting gBK might impede advancement to the terminal stage of SCLC via two pathways.

Histological analysis of human diced cartilage grafts.

BACKGROUND: Grafting the radix and dorsum of the nose has been controversial and the plastic surgery community has recently revisited diced cartilage grafts. Previous articles by the authors reported clinical failures of diced cartilage-Surgicel-wrapped grafts and the successful implementation of diced cartilage-fascia-wrapped grafts for rhinoplasty procedures. This article investigates four means of radix and dorsum augmentation, including banked and fresh septum. METHODS: Twenty-seven human specimens were taken from rhinoplasty patients: six failed diced cartilage-Surgicel-wrapped grafts, eight diced cartilage-fascia-wrapped grafts, and 13 specimens of fresh cartilage or previously placed cartilage grafts. All were stained with hematoxylin and eosin, Masson's trichrome, safranin-O, glial fibrillary acidic protein, and Evans' van Gieson stains and analyzed for chondrocyte viability, matrix proteins, and regenerative potential of the cartilage. RESULTS: The diced cartilage-Surgicel-wrapped grafts showed disorganized fibrosis, loss of nuclei in chondrocyte lacunae, and negative glial fibrillary acidic protein staining. All specimens contained remnants of Surgicel and evidence of foreign body reaction. The diced cartilage-fascia-wrapped grafts showed normal histologic characteristics, healthy cartilage pieces with viable chondrocytes in their lacunae, positive glial fibrillary acidic protein staining, and organized capsule of fibrosis surrounding the cartilage pieces. The fascia was viable and showed an organized architecture distinctly different from the disorganized scar tissue of the diced cartilage-Surgicel-wrapped grafts. CONCLUSIONS: Given the histologic findings, diced cartilage-Surgicel-wrapped grafts absorbed due to foreign body reaction. Diced cartilage-fascia wrapped grafts survived and demonstrated normal cartilage viability. According to the clinical performance and analysis of these grafts, diced cartilage-fascia is the most reliable way to augment the nasal dorsum and radix with autologous diced cartilage.