Good coronary collateral circulation is not associated with better prognosis in patients with chronic total occlusion, regardless of treatment strategy.

OBJECTIVE: This study aimed to assess the effects of coronary collateral circulation (CCC) on the prognosis of patients with chronic total occlusion (CTO) under different treatment strategies. METHODS: We analyzed a total of 1124 patients who were diagnosed with CTO and divided them into groups with good CCC (grade 2 to 3, n = 539) or poor CCC (grade 0 to 1, n = 531). The primary outcome was cardiac death during follow-up; the secondary outcome was major adverse cardiovascular events (MACEs). We also performed subgroup analyses in groups with and without CTO revascularization (CTO-R and CTO-NR, respectively), and sensitivity analyses excluding patients who received failed CTO-PCI to further investigate the effect of CCC. RESULTS: During a median follow-up duration of 23 months, we did not detect any significant differences between the good CCC group and the poor CCC group in terms of cardiac death (4.2% vs 4.1%; adjusted hazard ratio [HR], 1.01; 95% confidence interval [CI], 0.56-1.83; p = 0.970) and MACEs (23.6% vs 23.2%; adjusted HR, 1.07; 95% CI, 0.84-1.37; p = 0.590). Subgroup analyses according to CTO revascularization showed similar results. In addition, we observed no differences in sensitivity analyses when patients who received failed CTO-PCI were excluded. CONCLUSION: Good CCC was not associated with a lower risk of cardiac death or MACEs among patients with CTO, regardless of whether the patients received CTO revascularization treatment.

Effect of complete percutaneous revascularization on improving long-term outcomes of patients with chronic total occlusion and multi-vessel disease.

BACKGROUND: Limited data are available on the comparison of clinical outcomes of complete vs. incomplete percutaneous coronary intervention (PCI) for patients with chronic total occlusion (CTO) and multi-vessel disease (MVD). The study aimed to compare their clinical outcomes. METHODS: A total of 558 patients with CTO and MVD were divided into the optimal medical treatment (OMT) group ( n = 86), incomplete PCI group ( n = 327), and complete PCI group ( n = 145). Propensity score matching (PSM) was performed between the complete and incomplete PCI groups as sensitivity analysis. The primary outcome was defined as the occurrence of major adverse cardiovascular events (MACEs), and unstable angina was defined as the secondary outcome. RESULTS: At a median follow-up of 21 months, there were statistical differences among the OMT, incomplete PCI, and complete PCI groups in the rates of MACEs (43.0% [37/86] vs. 30.6% [100/327] vs. 20.0% [29/145], respectively, P = 0.016) and unstable angina (24.4% [21/86] vs. 19.3% [63/327] vs. 10.3% [15/145], respectively, P = 0.010). Complete PCI was associated with lower MACE compared with OMT (adjusted hazard ratio [HR] = 2.00; 95% confidence interval [CI] = 1.23-3.27; P = 0.005) or incomplete PCI (adjusted HR = 1.58; 95% CI = 1.04-2.39; P = 0.031). Sensitivity analysis of PSM showed similar results to the above on the rates of MACEs between complete PCI and incomplete PCI groups (20.5% [25/122] vs. 32.6% [62/190], respectively; adjusted HR = 0.55; 95% CI = 0.32-0.96; P = 0.035) and unstable angina (10.7% [13/122] vs. 20.5% [39/190], respectively; adjusted HR = 0.48; 95% CI = 0.24-0.99; P = 0.046). CONCLUSIONS: For treatment of CTO and MVD, complete PCI reduced the long-term risk of MACEs and unstable angina, as compared with incomplete PCI and OMT. Complete PCI in both CTO and non-CTO lesions can potentially improve the prognosis of patients with CTO and MVD.

Human adenovirus type 19 infection of corneal cells induces p38 MAPK-dependent interleukin-8 expression.

BACKGROUND: Human adenovirus type 19 (HAdV-19) is a major cause of epidemic keratoconjunctivitis, the only ocular adenoviral infection associated with prolonged corneal inflammation. In this study, we investigated the role of p38 mitogen-activated protein kinase (MAPK) in HAdV-19 infection, with particular attention to the role of p38 MAPK in the transcriptional control of interleukin-8 (IL-8), a chemokine previously shown to be central to the initiation of adenovirus keratitis. RESULTS: We found that infection of corneal cells with HAdV-19 led to activation of p38 MAPK and its downstream targets, HSP-27 and ATF-2, within 15 to 30 minutes post-infection. Infection also induced phosphorylation of IkappaB and NFkappaB in a p38 MAPK-dependent fashion. Furthermore, HAdV-19 induced an interaction between p38 MAPK and NFkappaB-p65, followed by nuclear translocation of activated NFkappaB-p65 and its binding to the IL-8 promoter. The interaction between p38 MAPK and NFkappaB-p65 was inhibited in concentration-dependent fashion by SB203580, a chemical inhibitor of p38 MAPK, but not by SP600125, an inhibitor of JNK - another MAPK implicated in chemokine expression by HAdV-19 infected cells. IL-8 gene expression in HAdV-19 infection was significantly reduced in the presence of sequence-specific p38 MAPK siRNA but not control siRNA. CONCLUSION: These results provide the first direct evidence for transcriptional regulation of IL-8 in HAdV-19 infected cells through the activation of the p38 MAPK signaling pathway. The p38 MAPK pathway may play a biologically important role in regulation of IL-8 gene expression in the adenovirus-infected cornea.

Functional role of JNK in neuritogenesis of PC12-N1 cells.

JNKs, also known as SAPKs, are activated in response to a wide variety of factors including growth factors, cytokines, UV radiation, and heat shock. In the rat pheochromocytoma PC12 cells, the JNK signaling pathway mediates diverse functions such as differentiation and apoptosis. We have previously shown that activated JNK is required for later stages of neuritogeneis induced by NGF in a variant PC12 cell line (N1). Here, the functional role of JNK in N1 cells is further investigated. We show that NGF treatment, which induces extensive neurite branching and cell soma enlargement in the N1 cells, stimulates a biphasic activation of JNK. The first phase of activation is rapid and transient, beginning at 15 min after NGF exposure and lasting approximately 45 min. The second phase of activation is sustained, beginning at 9-12 h of NGF treatment and lasting for at least 24 h. Similar biphasic pattern of JNK activation is also observed in the parental PC12 cells. Using the specific JNK inhibitor SP600125, we show that the biphasic activation is necessary for neurite outgrowth and branching, and that inhibition of either phase suppresses neuritogenesis in the N1 cells. These results suggest that dynamic JNK activation may play a key role in neurite outgrowth during neuronal development.

JNK regulates MCP-1 expression in adenovirus type 19-infected human corneal fibroblasts.

PURPOSE: Previous studies indicate that adenovirus type 19 (Ad19) infection of human corneal fibroblasts (HCFs) induces the expression of several proinflammatory mediators, including IL-8 and monocyte chemoattractant protein-1 (MCP-1), and that the tyrosine kinase c-Src and its downstream target, the mitogen-activated protein kinase ERK1/2, mediate IL-8 expression. In this context, the authors sought to investigate the potential role of another mitogen-activated protein kinase, c-Jun N-terminal kinase (JNK), in adenoviral ocular pathogenesis. METHODS: Ad19- and mock-infected HCFs were solubilized at various time points after infection, and cell lysates were subjected to SDS-PAGE followed by immunoblot analysis with a panel of antibodies against components of the MKK7/JNK/c-Jun pathway or immunoprecipitated for JNK assay. The induction of chemokine mRNA and protein was determined by real-time PCR and ELISA, respectively. RESULTS: Ad19 induced the phosphorylation of MKK7, JNK, and the downstream transcription factor c-Jun in HCFs at 15 and 30 minutes after infection. JNK activity was demonstrated at 30 minutes after infection using the GST-c-Jun fusion protein as a target substrate. SP600125, a specific pharmacologic inhibitor of JNK, blocked MCP-1 but not IL-8 mRNA and protein expression. Finally, PP2, a specific inhibitor of c-Src previously shown to inhibit the expression of both IL-8 and MCP-1 in Ad19-infected HCFs, also blocked JNK phosphorylation after infection. CONCLUSIONS: The MKK7/JNK/c-Jun cascade is rapidly activated and mediates MCP-1 expression in Ad19-infected HCFs. Furthermore, the activation of c-Src on Ad19 infection appears to regulate both the ERK and the JNK pathways.

Vitronectin: a possible determinant of adenovirus type 19 tropism for human corneal epithelium.

PURPOSE: Adenoviruses typically demonstrate specific tissue tropisms, as in the association of Ad19 with epidemic keratoconjunctivitis. We sought to determine factors that might influence the apparent tropism of Ad19 for the cornea. DESIGN: Laboratory investigation. METHODS: Adenovirus serotypes Ad2, 5, 9, 10, 11, 13, and 19 were compared for their capacity to replicate in human corneal epithelial cells (HCECs) in culture. Organotypically cultured human corneas were infected with Ad19 or Ad2, and viral titers were compared after 7 days. Replication of both viruses was compared in HCECs cultured on various extracellular matrices. Western blot analysis and immunohistochemistry were applied to human donor corneas and HCECs. RESULTS: One week after infection of HCEC monolayer cultures, Ad2 titers were significantly higher than any of the other viruses tested (P <.05). In organotypic corneal cultures, Ad19 titers were significantly higher than Ad2 (P = .0003). Ad2 replication in HCECs equaled or exceeded that of Ad19 on all extracellular matrices except vitronectin, where Ad2 replication was reduced and Ad19 replication enhanced (P <.0001). Vitronectin was detected by immunohistochemistry within the corneal epithelial basement membranes of human donor corneas. Increased alpha(v) integrin expression and greater tyrosine kinase phosphorylation in HCECs cultured on vitronectin were demonstrated by Western blot analysis. CONCLUSIONS: In vitro, vitronectin enhances growth of Ad19, possibly by up-regulation of receptor alpha(v) integrins and increased activity of tyrosine kinases necessary for adenoviral internalization. We hypothesize that differential tissue tropisms for adenoviruses may derive in part from tissue-specific extracellular matrix expression.

Kaposi sarcoma herpesvirus/human herpesvirus-8-negative effusion-based lymphoma: report of 3 cases and review of the literature.

BACKGROUND: Primary effusion lymphoma (PEL) is a rare subtype of large B-cell lymphoma that arises in body cavities without detectable tumor masses. PEL is universally associated with Kaposi sarcoma herpesvirus (KSHV)/human herpesvirus-8 (HHV8). Despite overlapping features, KSHV/HHV8-negative effusion-based lymphoma is a distinct entity from PEL. To date, 52 cases have been reported. The authors report 3 additional cases received in their laboratory from 2007 to 2012. METHODS: Clinical data, cytomorphologic features, and immunophenotypic features of the 3 cases were described and compared with those reported in the literature. RESULTS: The cells in HHV8-negative effusion lymphoma commonly revealed large cell, immunoblastic morphology and B-cell immunophenotype. The 3 cases demonstrated cytomorphologic and immunophenotypic variability. Cytomorphologically, 1 case contained large, highly atypical cells with a moderate amount of cytoplasm, round nucleus, coarsely granular chromatin, and a single macronucleolus. The other 2 cases had medium to large atypical cells with high nuclear-to-cytoplasmic ratios, slightly irregular to cleaved nuclei, and multiple conspicuous nucleoli. One case had a null phenotype with aberrant cytokeratin expression. B-cell phenotype was established by clonal immunoglobulin heavy-chain rearrangement using polymerase chain reaction, whereas the other 2 cases demonstrated a B-cell phenotype by flow cytometry and immunohistochemical staining. All 3 cases were negative for both HHV8 and Epstein-Barr virus. CONCLUSIONS: HHV8-negative effusion lymphoma exhibits clinical, cytomorphologic, and immunophenotypic variability. Cases with a null-phenotype can be particularly challenging. When effusion lymphoma is suspected, ancillary tests are helpful. Moreover, HHV8 detection is critical in differentiating PEL and HHV8-negative effusion lymphoma, because they have overlapping features yet different prognoses.

Differential roles of ERK and JNK in early and late stages of neuritogenesis: a study in a novel PC12 model system.

The rat pheochromocytoma PC12 cell line has been an invaluable model system for studying neuritogenesis. Nerve growth factor (NGF) elicits multiple aspects of neurite outgrowth in PC12 cells. It is therefore difficult to dissect and assign an individual signaling pathway to each stage of neuritogenesis. We have recently reported the isolation of a variant PC12 cell line, PC12-N1 (N1), which spontaneously extends neuritic processes and exhibits an increased sensitivity to NGF. Here, we show that, under different culture conditions, the cells display three distinct phases of neuritogenesis consisting of neurite initiation, rapid neurite elongation, and a maturation process characterized by the thickening of neurites and increase in cell soma sizes. We demonstrate that signaling through ERK, but not p38 or JNK, is required for the spontaneous neurite initiation and extension. Treatment with low concentrations of NGF induces rapid neurite elongation without affecting neurite branching and cell soma sizes. Such a rapid neurite outgrowth can be blocked by the inhibition of ERK, but not JNK, activities. In the presence of higher concentrations of NGF, the N1 cells undergo further differentiation with many characteristics of mature neurons in culture, e.g. larger cell soma and numerous branches/connections. This process can be completely blocked by inhibiting ERK or JNK activities using specific inhibitors. These results suggest that ERK and JNK signals play different roles in neuritogenesis, and that JNK activity is essential in the late stages of neuritogenesis. Furthermore, our results demonstrate that signaling dosage is important in the activation of a specific pathway, leading to distinctive biological outcomes.

Variant PC12 cell line that spontaneously differentiates and extends neuritic processes.

The rat pheochromocytoma PC12 cells differentiate into neuronal-like cells in response to treatment with neurotrophins. The cells have been extensively used for investigating neuronal differentiation and axonal growth. Here we report the isolation of a variant PC12 cell line, named PC12-N1, which spontaneously differentiates and extends neuritic processes. The PC12-N1 cells expressed many neuronal specific proteins, including the synaptosomal associated protein of 25 kDa (SNAP-25), synaptotagmin, and synaptobrevin (also known as VAMP). The cells also expressed neurofilament protein of 68 kDa, a marker for differentiated neurons. In addition to the spontaneous neurite outgrowth, the PC12-N1 cells showed a marked increase in neurite outgrowth upon treatment with nerve growth factor (NGF), basic fibroblast growth factor (bFGF), and cyclic AMP (cAMP). The activation of mitogen-activated protein (MAP) kinases was examined by immunoblot analysis using phospho-specific antibodies. No overactivation was observed with ERK1/2 or p38. However, the c-Jun N-terminal kinase JNK/SAPK was activated approximately 10-fold over the parental PC12 cells. These results suggest that activation of JNK/SAPK may be involved in the spontaneous neurite extension in the PC12-N1 cells. Moreover, the PC12-N1 cells may be used as a model for investigating molecular signaling mechanisms underlying neuronal differentiation and axonal outgrowth.

An immunohistochemical method that distinguishes free from complexed SNAP-25.

Soluble N-ethylmaleimide-sensitive fusion protein (NSF) attachment protein receptor (SNARE) complexes composed of target (t-) SNAREs syntaxin and SNAP-25 and vesicle SNARE synaptobrevin play an essential role in neurosecretion. It is hypothesized that a transient intermediate complex between the t-SNAREs is formed during the assembly of the ternary complex. The existence of the t-SNARE binary complexes in vivo, however, has not been demonstrated. By using an affinity absorption scheme with preformed syntaxin-SNAP-25 complexes, we isolated antibodies capable of distinguishing free SNAP-25 from those associated with syntaxin. By semiquantitative immunohistochemistry, we estimated that, in cultured cerebellar neurons, the majority of SNAP-25 existed as complexes. Compared with the cultured neurons, PC12 cells expressed significantly less syntaxin, and we found that SNAP-25 was primarily in free forms. In contrast, a PC12 line that stably expressed a recombinant syntaxin showed a marked increase in SNAP-25 complexes. By using fluorescence resonance energy transfer (FRET) techniques, we observed FRET between cyan fluorescence protein-syntaxin and yellow fluorescence protein-SNAP-25 fusion proteins expressed in COS-7 and PC12 cells, suggesting a physiological interaction between syntaxin and SNAP-25. Our results demonstrate that, unlike what was previously hypothesized, syntaxin and SNAP-25 exist preferably as stable binary complexes in neurons. These findings offer novel insight into the mechanisms underlying the initiation and regulation of SNARE complex assembly.