Involvement of the interferon-gamma-induced T cell-attracting chemokines,interferon-gamma-inducible 10-kd protein (CXCL10) and monokine induced by interferon-gamma (CXCL9), in the salivary gland lesions of patients with Sjögren's syndrome

OBJECTIVE: To elucidate the mechanism of the development of T cell infiltrates in the salivary glands of patients with Sj?gren's syndrome (SS), we studied T cell-attracting chemokines and their receptors. METHODS: The expression of the T cell-attracting chemokines, interferon-gamma (IFNgamma)-inducible 10-kd protein (IP-10; also called CXCL10), monokine induced by IFNgamma (Mig; also called CXCL9), and stromal cell-derived factor 1 (SDF-1; also called CXCL12), in salivary glands from SS patients was investigated by polymerase chain reaction-enzyme-linked immunosorbent assay (ELISA). Cells that produce chemokines and lymphocytes that express chemokine receptors were identified by immunohistochemistry. The production of IP-10 and Mig proteins by salivary epithelial cells in response to IFNgamma was determined by ELISA. RESULTS: Expression of IP-10 and Mig messenger RNA (mRNA) was significantly up-regulated in SS salivary glands compared with normal salivary glands (both P < 0.01). There was no significant difference in SDF-1 mRNA expression between the SS and normal salivary glands. IP-10 and Mig proteins were predominantly expressed in the ductal epithelium adjacent to lymphoid infiltrates. Most of the CD3+ infiltrating lymphocytes in dense periductal foci expressed CXCR3, the receptor for IP-10 and Mig. IFNgamma induced the production of high levels of IP-10 and Mig proteins from cultured SS salivary epithelial cells. CONCLUSION: These findings suggest that IFNgamma stimulates the production of IP-10 and Mig in the SS ductal epithelium, and that IP-10 and Mig are involved in the accumulation of T cell infiltrates in the SS salivary gland. Chemokines or chemokine receptors could be a rational new therapeutic target in SS.     

(23S)-25-Dehydro-1{alpha}-hydroxyvitamin D3-26,23-lactone, a vitamin D receptor antagonist that inhibits osteoclast formation and bone resorption in bone marrow cultures from patients with Paget's disease

Osteoclast (OCL) precursors from patients with Paget's disease (PD) and normal OCL precursors transduced with the measles virus nucleocapsid protein gene (MVNP) are hyperresponsive to 1alpha,25-dihydroxyvitamin D(3) [1alpha,25-(OH)(2)D(3)] and can form OCLs at physiologic concentrations of 1alpha,25-(OH)(2)D(3). This hyperresponsivity to 1alpha,25-(OH)(2)D(3) is due to increased expression of TATA box-associated factor II-17, a potential coactivator of the vitamin D receptor. Hyperresponsivity to 1alpha,25-(OH)(2)D(3) may permit OCL formation in PD patients with low levels of 1alpha,25-(OH)(2)D(3) and play a role in the pathogenesis of PD. Therefore, we tested the effects of a vitamin D antagonist, (23S)-25-dehydro-1alpha-hydroxyvitamin D(3)-26,23-lactone (TEI-9647), to determine its potential to inhibit the enhanced OCL formation and bone resorption seen in patients with PD. TEI-9647, by itself, was not a vitamin D receptor agonist and did not induce OCL formation in vitro, even at 10(-6) m. However, it dose-dependently (10(-10) m to 10(-6) m) inhibited osteoclast formation induced by concentrations of 1alpha,25-(OH)(2)D(3) (41 pg/ml, 10(-10) m) detected in PD patients by bone marrow cells of patients with PD and MVNP-transduced colony-forming unit-granulocyte macrophage (CFU-GM) cells, which form pagetic-like OCL. Moreover, bone resorption by OCLs derived from MVNP-transduced CFU-GM treated with 10(-9) m 1alpha,25-(OH)(2)D(3) was dose-dependently inhibited by TEI-9647 (10(-9) m to 10(-6) m). Furthermore, 10(-7) m TEI-9647 by itself did not cause 1alpha,25-(OH)(2)D(3)-dependent gene expression but almost completely suppressed expression of the TATA box-associated factor II-17 and 25-hydroxyvitamin D(3)-24-hydroxylase genes induced by 1alpha,25-(OH)(2)D(3) treatment of MVNP-transduced CFU-GM cells. These results demonstrate that TEI-9647 can suppress the excessive bone resorption and OCL formation seen in marrow cultures from patients with PD.     

Chronic ethanol consumption impairs cellular immune responses against HCV NS5 protein due to dendritic cell dysfunction

BACKGROUND & AIMS: Alcoholic patients with and without chronic liver disease have a high incidence of infection with hepatitis C virus (HCV). Long-term ethanol consumption in mice has been associated with a strikingly reduced CD8(+) cytotoxic T-lymphocyte (CTL) response to HCV nonstructural proteins following DNA-based immunization. This study evaluated the effect of ethanol on dendritic cells (DCs) as a mechanism(s) for reduced CTL activity. METHODS: Mice were fed an ethanol-containing or isocaloric pair-fed control diet for 8 weeks, followed by DC isolation from the spleen. DCs were evaluated with respect to endocytosis properties, cell surface markers, allostimulatory activity, and cytokine production following stimulation. Immune responses to HCV NS5 protein were generated by genetic immunization. Syngeneic transfer was used to determine if DC dysfunction contributed to abnormal cellular immune responses. RESULTS: Long-term ethanol exposure resulted in a reduced number of splenic DCs but did not alter endocytosis capacity. There was an increase in the myeloid and a reduction in the lymphoid DC population. Ethanol reduced expression of CD40 and CD86 costimulatory molecules on resting DCs, which was corrected following stimulation with lipopolysaccharide or poly I:C. There was impaired allostimulatory activity. Cytokine profiles of DCs isolated from ethanol-fed mice were characterized by enhanced interleukin (IL)-1beta and IL-10 and decreased tumor necrosis factor alpha, IL-12, interferon gamma, and IL-6 secretion. Impaired CTL responses to NS5 were corrected by syngeneic transfer of control DCs. CONCLUSIONS: Altered DC function is one of the major changes induced by long-term ethanol consumption, which subsequently impairs the cellular immune response necessary for viral clearance.     

Clinical significance of serum KL-6 and SP-D for the diagnosis and treatment of interstitial lung disease in patients with diffuse connective tissue disorders]

OBJECTIVE: To elucidate the clinical significance of serum KL-6 and SP-D for the diagnosis and treatment of interstitial lung disease in connective tissue disorders. METHODS: 139 patients with various connective tissue disorders were subjected for the study, which included 46 cases of rheumatoid arthritis, 43 cases of Sj?gren's syndrome, 16 cases of SLE, 10 cases of systemic sclerosis, 9 cases of polymyositis/dermatomyositis, 6 cases of vasculitis syndrome, 5 cases of Beh?et's disease and 4 cases of MCTD. Serum levels of KL-6 and SP-D were determined by enzyme-immunoassay. The sensitivity, specificity and accuracy of serum KL-6 and SP-D for the diagnosis of interstitial lung disease were compared with serum LDH. The relationship of serum KL-6 and SP-D levels with high resolution CT (HRCT) of the lung and Gallium scintigraphy findings was analyzed. In some cases, serum levels of the two markers were determined monthly in the course of the disease. RESULTS: When the serum levels of KL-6 and SP-D were measured simultaneously, the sensitivity to diagnose interstitial lung disease was 67.7%, the specificity was 98.1%, and the accuracy was 91.4%, while those of serum LDH were 45.2%, 88.9%, 79.1% respectively. In the patients with interstitial lung disease, those who had elevated serum levels of both KL-6 and SP-D showed parenchymal collapse opacity-dominant pattern in HRCT. On the other hand, the patients with interstitial lung disease who had normal levels of serum KL-6 and SP-D or had elevation either in KL-6 or SP-D levels showed ground glass opacity-dominant pattern in HRCT. There was no significant correlation between serum marker levels and Gallium scintigraphy findings. When serum KL-6 and SP-D were measured monthly, the levels of both markers changed more specifically and sensitively to the lung disease activity compared with serum LDH. CONCLUSIONS: Serum KL-6 and SP-D are more specific and useful markers for the diagnosis and evaluation of interstitial lung disease compared with serum LDH in connective tissue disorders.     

Clinical impact of interstitial pneumonia following surgery for lung cancer

BACKGROUND: Operative morbidity in patients with lung cancer associated with perioperative interstitial pneumonia (IP) has emerged as a serious problem. PATIENTS AND METHODS: We studied the clinical impact of perioperative related IP in 11 patients (IP group: 7 preoperative known, 4 acute onset) of 473 lung cancer patients who received a pulmonary resection. The IP group was compared to the remaining 462 patients (non-IP group). Demographic data, clinical presentation, and serum KL-6 levels were compared. RESULTS: There were no differences in age, gender, type of surgery, and pulmonary function except for % DLco between the non-IP and IP groups. The IP group showed a higher in-hospital mortality (n=2: 18.3%) than that of the non-IP group (n=3: 0.6%) (P<0.005). Seven patients with underlying IP with high KL-6 levels showed an uneventful recovery. Two patients with postoperative onset of acute IP had a fatal course associated with elevation of serum KL-6 levels. CONCLUSIONS: Postoperative development IP is a serious complication with high mortality, and serial measurement of KL-6 levels is useful to assess the activity of IP.     

Reduced expression and functional impairment of Toll-like receptor 2 on dendritic cells in chronic hepatitis C virus infection.

BACKGROUND: Dendritic cells (DCs) utilize Toll-like receptors (TLRs) to sense virus and initiate immune responses. We aimed at elucidating the roles of TLRs on DCs in hepatitis C virus (HCV) infection. METHODS: Monocyte-derived DCs were obtained from 32 healthy volunteers (HV) and 30 chronically HCV-infected patients (CH). TLR2, TLR3 and TLR4 expressions on immature DCs were quantified by real-time quantitative RT-PCR. We stimulated DCs with specific TLR ligands and examined DC maturation, cytokine production and ability to stimulate allogeneic CD4(+) T cells. RESULTS: TLR2 expression on immature DCs was lower in the CH group, whereas those of TLR3 or TLR4 were not different between the groups. Each TLR ligand induced DC maturation and stimulated them to release comparable levels of IL-12p70, IL-6, IL-10, TNF-alpha and IFN-beta between the groups. TLR2 and TLR4 ligands enhanced DC ability to stimulate T cell proliferation, with the degree due to the TLR2 ligand being lower in the CH group. CONCLUSIONS: In HCV infection, the TLR2 expression on DCs is reduced and TLR2-stimulated DCs show lesser ability to proliferate T cells than healthy counterparts, suggesting that the TLR2 system is involved in HCV-induced immunopathogenesis.     

Pharmacological Profile of U‐37883A,a Channel Blocker of Smooth Muscle‐Type ATP‐Sensitive K+ Channels

U‐37883A (PNU‐37883A, guanidine; 4‐morpholinecarboximidine‐N‐1‐adamantyl‐N′‐cyclohexyl hydrochloride) was originally developed as a potential diuretic with specific binding in kidney and vascular smooth muscle rather than in brain or pancreatic β cells. U‐37883A inhibits ATP‐sensitive K⁺ channels (K_ATP channels) in vascular smooth muscle at submicromolar concentrations whilst even at high concentrations (≥10 μM) it has no inhibitory effect at pancreatic, cardiac or skeletal K_ATP channels. Thus, it is generally thought that U‐37883A is a selective inhibitor of vascular smooth muscle K_ATP channels. Approximately one decade ago, K_ATP channels were cloned and found to consist of at least two subunits: an inwardly‐rectifying K⁺ channel six family (K_ir6.x; K_ir6.1 and K_ir6.2) which forms the ion conducting pore and a modulatory sulphonylurea receptor (SUR.x; SUR1, SUR2A, and SUR2B) that accounts for several pharmacological properties. It is generally believed that different combinations of K_ir6.x and SUR.x determine the molecular properties of K_ATP channels. Thus, K_ir6.2/SUR1 channel represents the pancreatic β‐cell K_ATP channel, K_ir6.2/SUR2A channel is thought to represent the cardiac K_ATP channel, whereas K_ir6.1/SUR2B channel is likely to represent the vascular smooth muscle K_ATP channel. Recent molecular studies have shown that U‐37883A selectively suppresses the activity of recombinant K_ATP channels which contain K_ir6.1 subunits in the channel pore unit. It was thus thought that U‐37883A was a selective pharmacological tool which could be used to investigate the activity of vascular smooth muscle K_ATP channels. However, due to its multiple pharmacological actions on several ion channels and poor tissue selectivity, U‐37883A should not be viewed as a selective blocker of smooth muscle K_ATP channels.     

Effectiveness of real-time virtual sonography-guided radiofrequency ablation treatment for patients with hepatocellular carcinomas.

Aim: Real-time virtual sonography (RVS) can synchronize B-mode ultrasound (US) images with multiplanar reconstruction (MPR)-computed tomography (CT) images on the same screen in real time. The purpose of this study was to evaluate the effectiveness of RVS for radiofrequency ablation therapy (RFA) of hepatocellular carcinomas (HCC) in which it was difficult to identify contours or margins by B-mode US. Methods: Sixty-three consecutive patients with a solitary HCC of less than 3.5 cm in diameter were enrolled in this study. Thirty-nine patients with HCC clearly detectable by B-mode US underwent conventional RFA, while the remaining 24 with obscure tumor lesions underwent RVS-guided RFA. A follow-up study of RFA treatment was performed every 3 months using enhanced CT imaging of the arterial and portal phase (at least 24 months). The accuracy of needle insertion was confirmed by measuring the gap between the needle insertion line and the center of the tumor from MPR-CT images. Results: The local recurrence rate of the RVS-guided RFA group was similar to that of the conventional RFA group (8.3% vs 7.7%), despite the difficulty of detecting tumor lesions in the former group. The mean gap between the needle insertion line and the center of the tumor was 1.6 mm (0-3.2 mm) in eight patients treated with RVS-guided RFA. Conclusion: RVS-guided RFA can be useful for treating HCC that are difficult to detect by B-mode US.     

Erythropoiesis failure due to RPS19 deficiency is independent of an activated Tp53 response in a zebrafish model of Diamond-Blackfan anaemia

Diamond-Blackfan anaemia (DBA) is a cancer-prone genetic disorder characterized by pure red-cell aplasia and associated physical deformities. The ribosomal protein S19 gene (RPS19) is the most frequently mutated gene in DBA (~25%). TP53-mediated cell cycle arrest and/or apoptosis in erythroid cells have been suggested to be major factors for DBA development, but it is not clear why mutations in the ubiquitously expressed RPS19 gene specifically affect erythropoiesis. Previously, we showed that RPS19 deficiency in zebrafish recapitulates the erythropoietic and developmental phenotypes of DBA, including defective erythropoiesis with severe anaemia. In this study, we analysed the simultaneous loss-of-function of RPS19 and Tp53 in zebrafish to investigate the role of Tp53 in the erythroid and morphological defects associated with RPS19 deficiency. Co-inhibition of Tp53 activity rescued the morphological abnormalities, but did not alleviate erythroid aplasia in RPS19-deficient zebrafish. In addition, knockdown of two other RP genes, rps3a and rpl36a, which result in severe morphological abnormalities but only mild erythroid defects, also elicited an activated Tp53 response. These results suggest that a Tp53-independent but RPS19-dependent pathway could be responsible for defective erythropoiesis in RPS19-deficient zebrafish.     

Hemi-clamshell approach for advanced primary lung cancer

BACKGROUND: The hemi-clamshell approach provides a wide anterior view of the mediastinum, apical dome, and cervicothoracic area. However, only a few reports have been made regarding this technique. METHODS: The hemi-clamshell approach was used in 24 patients, of whom 5 had a Pancoast tumor, 15 had mediastinal involvement, and 4 underwent mediastinal lymphadenopathy for left-sided lung cancer. Twenty-one of the patients received preoperative therapy. RESULTS: Twenty-one operations were complete resections. In addition, 12 patients received cardio-vascular reconstruction and 5 a first rib resection. Postoperative major morbidity was 21 % (6/24) and mortality was 4.2 % (1/24). Nine patients died of systemic tumor relapse and 14 patients were alive after a median follow-up of 24 months (range 3 - 68 months) following the initial therapy. The 5-year survival rate of patients with mediastinal involvement was 37 % and that of 13 patients with postoperative stage I or II was 35 %. CONCLUSIONS: The hemi-clamshell approach provides a wide exposure allowing a safe and complete removal of lung cancer that involves the mediastinum and apical thoracic dome, leading to a better long-term survival rate for patients with this disease.