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BACKGROUND: In a phase III trial in patients with advanced, well-differentiated, progressive pancreatic neuroendocrine tumors, sunitinib 37.5 mg/day improved investigator-assessed progression-free survival (PFS) versus placebo (11.4 vs. 5.5 months; HR, 0.42; P < 0.001). Here, we present PFS using retrospective blinded independent central review (BICR) and final median overall survival (OS), including an assessment highlighting the impact of patient crossover from placebo to sunitinib.PATIENTS AND METHODS: In this randomized, double-blind, placebo-controlled study, cross-sectional imaging from patients was evaluated retrospectively by blinded third-party radiologists using a 2-reader, 2-time-point lock, followed by a sequential locked-read, batch-mode paradigm. OS was summarized using the Kaplan-Meier method and Cox proportional hazards model. Crossover-adjusted OS effect was derived using rank-preserving structural failure time (RPSFT) analyses.RESULTS: Of 171 randomized patients (sunitinib, n = 86; placebo, n = 85), 160 (94%) had complete scan sets/time points. By BICR, median (95% confidence interval [CI]) PFS was 12.6 (11.1-20.6) months for sunitinib and 5.8 (3.8-7.2) months for placebo (HR, 0.32; 95% CI 0.18-0.55; P = 0.000015). Five years after study closure, median (95%CI) OS was 38.6 (25.6-56.4) months for sunitinib and 29.1 (16.4-36.8) months for placebo (HR, 0.73; 95% CI 0.50-1.06; P = 0.094), with 69% of placebo patients having crossed over to sunitinib. RPSFT analysis confirmed an OS benefit for sunitinib.CONCLUSIONS: BICR confirmed the doubling of PFS with sunitinib compared with placebo. Although the observed median OS improved by nearly 10 months, the effect estimate did not reach statistical significance, potentially due to crossover from placebo to sunitinib.TRIAL REGISTRATION NUMBER: NCT00428597.

Background: Free sensate thoracodorsal artery perforator flaps that include the posterior divisions of the lateral cutaneous branches of the intercostal nerves have been described. The authors used preoperative color Doppler sonography to identify the nerves and demonstrate its clinical value. Methods: Fourteen free sensate thoracodorsal artery perforator flaps were collected. Preoperative color Doppler sonography was used to identify the locations of thoracodorsal artery perforators and the courses of the posterior divisions of the lateral cutaneous branches of the intercostal neurovascular bundles. These posterior divisions were preserved on flaps and classified into three types. Type A and B nerves sprouted cutaneous perforating fascicles over the lateral region of the latissimus dorsi muscle. Type C nerves went through the region without any dominant perforating fascicle. Results: Twenty-one nerves were mapped, and 24 were found during surgery. The sensitivity of preoperative color Doppler sonography was 87.5 percent. Of the 24 nerves, nine were type A (37.5 percent), 12 were type B (50 percent), and three were type C (12.5 percent). Ten of the 14 patients (sensate group) showed better tactile recovery at both the center and the periphery of flap than the other 10 patients who underwent reconstruction with nonsensate flaps. Conclusions: Preoperative color Doppler sonography is an indispensable tool for sensate thoracodorsal artery perforator flaps in locating the perforators and mapping the posterior divisions of the lateral cutaneous branches of the intercostal nerves. The information can be used to design and harvest sensate thoracodorsal artery perforator flaps, which are associated with earlier and better tactile recovery.

Background: The thoracodorsal artery perforator flap is a versatile flap for resurfacing soft-tissue defects. However, it is too bulky for resurfacing shallow defects. The authors used preoperative color Doppler sonography to identify the suprafascial courses of skin perforators to facilitate the design and thinning procedures of thoracodorsal artery perforator flaps. Methods: Thin thoracodorsal artery perforator flaps were designed in 29 patients. Preoperative color Doppler sonography was used to identify the penetrating points and suprafascial courses of skin perforators. According to the different orientations of suprafascial courses, skin perforators could be classified into three types. The designs and thinning procedures of flaps relied on the findings of preoperative color Doppler sonography. Results: Fifty-eight ideal perforators were marked on the 29 patients. Intraoperative dissections proved that 55 perforators were thoracodorsal artery skin perforators, and three perforators were intercostal artery perforators. In one patient, no skin perforator derived from the descending branch of the thoracodorsal artery was found during intraoperative dissection (3.4 percent). Suprafascial courses could be demonstrated by preoperative color Doppler sonography in 54 of the 55 thoracodorsal artery skin perforators. Nineteen were type 1 perforators (35.2 percent), 26 were type 2 perforators (48.1 percent), and nine were type 3 perforators (16.7 percent). Complications were encountered in six patients (20.7 percent), all of whom recovered well. Conclusions: The suprafascial courses of skin perforators facilitate the design and thinning procedures of thoracodorsal artery perforator flaps. They improve the survivability of flaps and make thinning procedures more secure and efficient.

1. Short-wave diathermy (SWD) is a form of radiofrequency radiation that is used therapeutically by physiotherapists. The cellular mechanisms of SWD are unclear. The present study was performed to explore the effect of different conditions of short-wave exposure on the proliferation of cultured human chondrocytes. 2. Cells exposed to short waves once per day for seven consecutive days exhibited a significant increase in proliferation by 42% compared with the control cells. In cells that were treated with short waves twice per day for seven consecutive days, or only once on Day 1 and then examined for proliferation on Day 7, cell proliferation was greater than the control cells by 40% and 30%, respectively. 3. Given the importance of mitogen-activated protein kinases (MAPK) in the proliferation of different cell types, efforts were extended to explore the role of three major types of MAPK; that is, extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal protein kinase (JNK) and p38. 4. It was found that the level of phosphorylated ERK (phospho-ERK 1 and ERK 2) increased significantly within 5-120 min following consecutive exposure to short waves for 7 days. Exposure to short waves failed to alter the intensity of phosphorylated JNK and p38 within 0-240 min. 5. Cells were exposed to short waves once for seven consecutive days in the presence of 0, 10 micromol/L, 20 micromol/L or 50 micromol/L PD98059 (an ERK inhibitor). PD98059 totally inhibited short waves-induced enhancement of proliferation without altering normal control viability. In the presence of short waves and PD98059, the cell viability was lower than the normal control. Together, the data suggest that short waves could increase proliferation in human chondrocytes through activation of the ERK pathway, which is also involved in maintaining normal cell proliferation under physiological conditions.

The effect of anandamide on human osteoblasts is unclear. This study examined the effect of anandamide on viability, apoptosis, mitogen-activated protein kinases (MAPKs) and Ca2+ levels in MG63 osteosarcoma cells. Anandamide at 50-200 microM decreased cell viability via apoptosis as demonstrated by propidium iodide staining and activation of caspase-3. Immunoblotting suggested that anandamide induced expression of ERK, JNK and p38 MAPK. Anandamide-induced cell death and apoptosis were reversed by SB203580, but not by PD98059 and SP600125, suggesting that anandamide's action was via p38 MAPK, but not via ERK and JNK. Anandamide at 1-100 microM induced [Ca2+]i increases. Removal of extracellular Ca2+ decreased the anandamide response, indicating that anandamide induced Ca2+ influx and Ca2+ release. Chelation of intracellular Ca2+ with BAPTA reversed anandamide-induced cell death and p38 MAPK phosphorylation. Collectively, in MG63 cells, anandamide induced [Ca2+]i increases which evoked p38 MAPK phosphorylation. This p38 MAPK phosphorylation subsequently activated caspase-3 leading to apoptosis.

The effects of econazole, an antifungal drug applied for treatment of keratitis and mycotic corneal ulcer, on cytosolic-free Ca(2+) concentrations ([Ca(2+)](i)) and viability of corneal cells was examined by using SIRC rabbit corneal epithelial cells as model. [Ca(2+)](i) and cell viability were measured by using the fluorescent dyes fura-2 and WST-1, respectively. Econazole at concentrations > or = 1 microM increased [Ca(2+)](i) in a concentration-dependent manner. The Ca(2+) signal was reduced partly by removing extracellular Ca(2+). The econazole-induced Ca(2+) influx was insensitive to L-type Ca(2+) channel blockers and protein kinase C modulators. In Ca(2+)-free medium, after pretreatment with 20 microM econazole, [Ca(2+)](i) rises induced by 1 microM thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor) were abolished. Conversely, thapsigargin pretreatment also abolished econazole-induced [Ca(2+)](i) rises. Inhibition of phospholipase C with 2 microM U73122 did not change econazole-induced [Ca(2+)](i) rises. At concentrations between 10 and 80 microM, econazole killed cells in a concentration-dependent manner. The cytotoxic effect of 20 microM econazole was not reversed by prechelating cytosolic Ca(2+) with BAPTA. This shows that in SIRC cells econazole induces [Ca(2+)](i) rises by causing Ca(2+) release from the endoplasmic reticulum and Ca(2+) influx from unknown pathways. Econazole-caused cytotoxicity was independent from a preceding [Ca(2+)](i) rise.

The effect of the carcinogen safrole on intracellular Ca2+ movement has not been explored in osteoblast-like cells. This study examined whether safrole could alter Ca2+ handling and viability in MG63 human osteosarcoma cells. Cytosolic free Ca2+ levels ([Ca2+]i) in populations of cells were measured using fura-2 as a fluorescent Ca2+ probe. Safrole at concentrations above 130 microM increased [Ca2+]i in a concentration-dependent manner with an EC50 value of 450 microM. The Ca2+ signal was reduced by 30% by removing extracellular Ca2+. Addition of Ca2+ after safrole had depleted intracellular Ca2+ induced Ca2+ influx, suggesting that safrole caused Ca2+ entry. In Ca2+-free medium, after pretreatment with 650 microM safrole, 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) failed to release more Ca2+; and pretreatment with thapsigargin inhibited most of the safrole-induced [Ca2+]i increases. Inhibition of phospholipase C with U73122 did not affect safrole-induced Ca2+ release; whereas activation of protein kinase C with phorbol ester enhanced safrole-induced [Ca2+]i increase. Trypan exclusion assays revealed that incubation with 65 microM safrole for 30 min did not kill cells, but incubation with 650 microM safrole for 10-30 min nearly killed all cells. Flow cytometry demonstrated that safrole evoked apoptosis in a concentration-dependent manner. Safrole-induced cytotoxicity was not reversed by chelation of Ca2+ with BAPTA. Collectively, the data suggest that in MG63 cells, safrole induced a [Ca2+]i increase by causing Ca2+ release mainly from the endoplasmic reticulum in a phospholipase C-independent manner. The safrole response involved Ca2+ influx and is modulated by protein kinase C. Furthermore, safrole can cause apoptosis in a Ca2+-independent manner.

Background Early colloid resuscitation in major burn patients has been stopped because of its deteriorating effect on thermal injury-induced vascular hyperpermeability. We hypothesized that inhibition of inducible nitric oxide synthase (iNOS) to stabilize endothelial permeability and to retain colloid solution in the vascular space will reverse its effect on lung damage. Study design In experiment 1, specific pathogen free rats underwent 35% total-body surface area burn or sham burn and were given equal volumes (7.5 mL/kg) of normal saline or albumin from femoral veins for fluid resuscitation immediately after burn. In experiment 2, S-methylisothiourea (SMT, 7.5 mg/kg, IP) was given immediately after burn to rats from different groups, as in experiment 1. At 8 hours after burn, blood was assayed for peroxynitrite-mediated dihydrorhodamine 123 (DHR 123) oxidation, and lung tissues were harvested for myeloperoxidase (MPO) determination and histologic studies. Pulmonary microvascular dysfunction was quantified by measuring the extravasations of Evans blue dye. Results Blood peroxynitrite level and iNOS expression, MPO activity, permeability, and inflammatory cell infiltration of lungs were significantly induced after thermal injury. Albumin resuscitation after burn without iNOS inhibition enhanced thermal injury-induced lung damage with 10%, 14%, and 5% increases in blood DHR oxidation level, lung MPO activity, and lung permeability, respectively, compared with saline injection. In contrast, burn + SMT rats with albumin injection showed significant, 23%, 37%, and 20%, decreases, respectively, in blood DHR 123 oxidation level, lung MPO activity, and lung permeability compared with burn + SMT + saline rats. Conclusions Thermal injury induced lung damage. Restoration of extracellular fluid in early burn shock with albumin markedly augmented the lung neutrophil deposition, lung permeability increase, and blood peroxynitrite level. Inhibition of iNOS before albumin supplementation reversed its damaging effects on thermal injury-induced lung dysfunction to beneficial ones.

The effect of miconazole, an anti-fungal drug, on cytoplasmic free Ca 2+ concentrations ([Ca 2+ ] i ) in human osteosarcoma cells (MG63) was explored by using the Ca 2+ -sensitive dye fura-2. Miconazole acted in a concentration-dependent manner with an EC 50 of 75 μM. The Ca 2+ signal comprised a gradual rise and a sustained elevation. Removal of extracellular Ca 2+ reduced 50% of the signal. In Ca 2+ -free medium, the [Ca 2+ ] i rise induced by 1 μM thapsigargin (an endoplasmic reticulum Ca 2+ pump inhibitor) was completely inhibited by pretreatment with 20 μM miconazole. Pretreatment with thapsigargin partly inhibited miconazole-induced Ca 2+ release. The miconazole-induced Ca 2+ release was not changed by inhibition of phospholipase C with 2 μM U73122. By using tetrazolium as a fluorescent probe, it was shown that 10–100 μM miconazole decreased cell proliferation rate in a concentration-dependent manner. Collectively, this study shows that miconazole induces [Ca 2+ ] i rises in human osteosarcoma cells via releasing Ca 2+ mainly from the endoplasmic reticulum in a manner independent of phospholipase C activity, and by causing Ca 2+ influx. Furthermore, miconazole may be cytotoxic to the cells at higher concentrations.

The differences in the intracellular Ca2+ responses to hormones in platelets from systemic lupus erythematosus (SLE) patients compared to normal humans have not been explored. This study examined the Ca2+ signaling and density of platelets in normal, inactive and active SLE patients. The platelet number per µl in inactive and normal groups did not differ, whereas the number in active SLE patients was smaller than the other two groups by 60%. The intracellular free Ca2+ levels ([Ca2+]i) in response to stimulation of four endogenous Ca2+ mobilizing hormones, 100 µM arachidonic acid (AA), 10 µM ADP, 10 nM platelet activation factor (PAF) and 1 µM thrombin, were investigated using the Ca2+-sensitive fluorescent dye, fura-2. The AA-induced [Ca2+]i rises in normal and inactive groups were similar. In contrast, the AA-induced [Ca2+]i rises in the active SLE group were significantly smaller than in the normal and inactive groups. The defect in the AA-induced [Ca2+]i rises in active SLE groups appears to be caused by defective Ca2+ influx and Ca2+ releasing pathways because the AA-induced responses were not altered by removal of extracellular Ca2+, whereas the AA-induced responses in normal and inactive SLE groups were reduced by removal of extracellular Ca2+, and the AA-induced Ca2+ release was smaller in the active SLE group. PAF, ADP and thrombin all induced [Ca2+]i rises in the three groups, but no significant differences were found among the three groups. Together, the results indicate that cell density and Ca2+ signaling in platelets from active SLE patients are altered in response to particular stimulators. In these regards, platelets from inactive SLE patients appear to be similar to those from normal humans.

Objective: To determine whether inhibition of inducible nitric oxide synthase to stabilize endothelial permeability and to retain hypertonic saline in the vascular space will ameliorate burn-induced gut barrier dysfunction. Design: Prospective, experimental study. Setting: Research laboratory at a university hospital. Subjects: Thermal injury models in the rat. Interventions: In experiment 1, specific pathogen free rats underwent 3% total body surface area burn or sham burn and were given 7.5 mL/kg hypertonic saline (7.5% NaCl), 7.5 mg/kg saline, or 50 mL/kg saline (nearly equal sodium load with hypertonic saline) in the right femoral vein for 15 mins for fluid resuscitation at 0, 4, or 8 hrs after burn. In experiment 2, S-methylisothiourea (7.5 mg/kg, intraperitoneally), a specific inducible nitric oxide synthase inhibitor, was given immediately after burn to rats from different groups as in experiment 1. At 24 hrs after burn, the intestinal mucosa was assayed for myeloperoxidase activity and lipid peroxidation, the distribution of fluorescein isothiocyanate-dextran across the lumen of the small intestine was determined, and bacterial translocation to the mesenteric lymph nodes and ileum histology were also examined. Measurements and Main Results: Burn induced significant increases in intestinal mucosa inducible nitric oxide synthase expression, myeloperoxidase activity, lipid peroxidation, intestinal permeability, bacterial translocation to mesenteric lymph nodes, and villi sloughing in rats. Hypertonic saline administration at 0 or 4 hrs after burn worsened intestinal mucosa lipid peroxidation, neutrophil sequestration, intestinal permeability, and villi sloughing compared with those of burn + 7.5 mg/kg saline and burn + 50 mL/kg saline rats. To the contrary, burn + S-methylisothiourea rats with hypertonic saline injection at 4 or 8 hrs after burn showed an improvement of gut barrier function compared with burn + S-methylisothiourea + 7.5 mg/kg saline and burn + S-methylisothiourea + 50 mL/kg saline rats. Administration of hypertonic saline at 8 hrs after burn and S-methylisothiourea injection also significantly attenuated the bacterial translocation to mesenteric lymph nodes and villi sloughing. Conclusions: Using hypertonic saline as a resuscitation fluid in early burn shock markedly augmented the thermal injury-induced intestinal mucosa neutrophil deposition, lipid peroxidation, and intestinal hyperpermeability. Inhibition of inducible nitric oxide synthase not only significantly attenuated neutrophil deposition and mucosa lipid peroxidation but also reversed the deteriorating effects of hypertonic saline on thermal injury-induced gut barrier dysfunction and bacterial translocation.

1. In human MG63 osteosarcoma cells, the effect of calmidazolium on [Ca 2 + ] i and proliferation was explored using fura-2 and ELISA, respectively. 2. Calmidazolium, at concentrations greater than 0.1 μmol/L, caused a rapid increase in [Ca 2 + ] i in a concentration-dependent manner (EC 5 0 = 0.5 μmol/L). The calmidazolium-induced [Ca 2 + ] i increase was reduced by 66% by removal of extracellular Ca 2 + . In Ca 2 + -free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2 + -ATPase, caused a monophasic increase in [Ca 2 + ] i , after which the effect of calmidazolium to increase [Ca 2 + ] i was completely inhibited. U73122, an inhibitor of phospholipase C (PLC), abolished histamine (but not calmidazolium)-induced increases in [Ca 2 + ] i . Pretreatment with phorbol 12-myristate 13-acetate to activate protein kinase C inhibited the calmidazolium-induced increase in [Ca 2 + ] i in Ca 2 + -containing medium by 47%. 3. Separately, it was found that overnight treatment with 2-10 μmol/L calmidazolium inhibited cell proliferation in a concentration-dependent manner. 4. These results suggest that calmidazolium increases [Ca 2 + ] i by stimulating extracellular Ca 2 + influx and also by causing release of intracellular Ca 2 + from the endoplasmic reticulum in a PLC-independent manner. Calmidazolium may be cytotoxic to osteosarcoma cells.

The effect of the antidepressant nortriptyline, on bone cells is unknown. In human osteosarcoma MG63 cells, the effect of nortriptyline on intracellular Ca2+ concentration ([Ca2+]i) and proliferation was measured by using fura-2 and tetrazolium, respectively. Nortriptyline (> or = 10 microM) caused a [Ca2+]i rise in a concentration-dependent manner (EC50 = 200 microM). Nortriptyline-induced [Ca2+]i rise was prevented by 60% by removal of extracellular Ca2+ but was not altered by voltage-gated Ca2+ channel blockers. In Ca2+ -free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca2+ -ATPase, caused a monophasic [Ca2+]i rise, after which the increasing effect of nortriptyline on [Ca2+]i was abolished; also, pretreatment with nortriptyline abolished thapsigargin-induced [Ca2+]i increase. U73122, an inhibitor of phospholipase C, did not affect nortriptyline-induced [Ca2+]i rise; however, activation of protein kinase C decrease nortriptyline-induced [Ca2+]i rise by 32%. Overnight incubation with 50 and 100 microM nortriptyline killed 78% and 97% of cells, respectively; while 10 microM nortriptyline had no effect. These data suggest that nortriptyline rapidly increases [Ca2+]i in human osteosarcoma cells by stimulating both extracellular Ca2+ influx and intracellular Ca2+ release, and is cytotoxic at high concentrations.

In human neuroblastoma IMR32 cells, the effect of the anti-depressant maprotiline on baseline intracellular Ca 2+ concentrations ([Ca 2+ ] i ) was explored by using the Ca 2+ -sensitive probe fura-2. Maprotiline at concentrations greater than 100 μM caused a rapid rise in [Ca 2+ ] i in a concentration-dependent manner (EC 50 = 200 μM). Maprotiline-induced [Ca 2+ ] i rise was reduced by 50% by removal of extracellular Ca 2+ . Maprotiline-induced [Ca 2+ ] i rises were inhibited by half by nifedipine, but was unaffected by verapamil or diiltiazem. In Ca 2+ -free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2+ -ATPase, caused a monophasic [Ca 2+ ] i rise, after which the increasing effect of maprotiline on [Ca 2+ ] i was abolished. U73122, an inhibitor of phospholipase C, did not affect maprotiline-induced [Ca 2+ ] i rises. These findings suggest that in human neuroblastoma cells, maprotiline increases [Ca 2+ ] i by stimulating extracellular Ca 2+ influx and also by causing intracellular Ca 2+ release from the endoplasmic reticulum via a phospholiase C-independent manner.

Addition of cetuximab showed trend, but not significantly, to improve the therapeutic effects of gemcitabine and oxaliplatin combination for advanced biliary tract cancer in current randomized phase II trial. The trend of improvement did not correlate with KRAS mutation status. Biomarkers finding for selecting patients who will benefit most from anti-EGFR targeted therapy is warranted. Background Previous clinical trials have not proved that adding epidermal growth factor receptor inhibitors to chemotherapy confers a survival benefit for patients with advanced biliary tract cancer (ABTC). Whether the KRAS mutation status of tumor cells confounded the results of past studies is unknown. Patients and methods ABTC patients stratified by KRAS status, Eastern Cooperative Oncology Group performance status, and primary tumor location were randomized 1 : 1 to receive GEMOX (800 mg/m2 gemcitabine and 85 mg/m2 oxaliplatin) or C-GEMOX (500 mg/m2 cetuximab plus GEMOX) every 2 weeks. The primary end point was objective response rate (ORR). Results The study enrolled 122 patients between December 2010 and May 2012 (62 treated with C-GEMOX and 60 with GEMOX). Compared with GEMOX alone, C-GEMOX was associated with trend to better ORR (27% versus 15%; P = 0.12) and progression-free survival (PFS, 6.7 versus 4.1 months; P = 0.05), but not overall survival (OS, 10.6 versus 9.8 months; P = 0.91). KRAS mutations, which were detected in 36% of tumor samples, did not affect the trends of difference in ORR and PFS between C-GEMOX and GEMOX. The two treatment arms had similar adverse events, except that more patients had skin rashes, allergic reactions, and neutropenia in the C-GEMOX arm. Of patients with C-GEMOX, the presence of a grade 2 or 3 skin rash was associated with significantly better ORR, PFS, and OS. Conclusions Addition of cetuximab did not significantly improve the ORR of GEMOX chemotherapy in ABTC, although a trend of PFS improvement was observed. The trend of improvement did not correlate with KRAS mutation status. Clinical Trials number This study is registered at ClinicalTrials.gov (NCT01267344). All patients gave written informed consent.

This study was designed to investigate the role of NO and effect of iNOS inhibitor on the lung neutrophil deposition and damage after burn. In Experiment 1, specific pathogen-free (SPF) Sprague-Dawley rats underwent 35% total body surface area (TBSA) burn. On the 4th, 8th, 16th, and 24th h after burn, blood was collected for peroxynitrite-mediated dihydrorhodamine 123 (DHR 123) oxidation assay, and lung tissues were harvested for myeloperoxidase (MPO) test and histologic study. Pulmonary microvascular dysfunction was quantitated by measuring the extravasation of Evans blue dye (EBD). In Experiment 2, S-methylisothiourea (SMT) was given (7.5 mg/kg, intraperitoneal immediately post-burn) to suppress iNOS activity. On the 8th h after burn, the effect of SMT on blood DHR 123 oxidation, lung MPO, lung damage, and lung iNOS expression were evaluated. Lung MPO activity increased up to a maximum of 2-fold 8 h after burn. Blood DHR 123 oxidation increased up to a maximum of 2-fold 8 h after burn. Lung permeability increased up to a maximum of 2.5-fold 4 h after burn. SMT significantly decreased lung MPO activity, blood DHR 123 oxidation, and lung permeability by 31%, 41%, and 54%, respectively. SMT markedly decreased the thermal injury-induced perivascular and interstitial inflammatory cell infiltration and iNOS staining in bronchiolar epithelium, endothelial cells, and perivascular and interstitial inflammatory cells. In conclusion, thermal injury induces blood DHR 123 oxidation, lung neutrophil deposition, lung iNOS expression, and lung damage. Peroxynitrite might play an important role in thermal injury-induced lung neutrophil deposition and damage. Specific inhibition of lung iNOS expression and blood DHR 123 oxidation with SMT on thermal injury not only attenuated the lung neutrophil deposition, but also reduced lung damage.

Expression of the p16 gene from 30 malignant skin tumors has been surveyed by immunohistochemical assay. Gene point mutations were detected by DNA direct sequencing and the mRNA level of gene expression was measured by RT-PCR. A silent point mutation of the p16 gene was found in only one patient. However, loss of expression of the p16 gene was noticed in 23 of 29 samples (79.3%). Correlation between loss of expression of the p16 gene and metastasis is significant (p = 0.0036). These findings suggest that loss of expression of the p16 gene may play a critical role in tumor progression of malignant skin tumors.

Extraspinal musculoskeletal tuberculosis (TB) was an uncommon manifestation with a frequency about 1 to 2% in all TB patients. According to the report, tuberculous arthritis and osteomyelitis remained the two leading etiologies, while others constituted less than 2% of all extraspinal musculoskeletal tuberculous infections; less than 0.04% of those who suffered from TB were represented as extraspinal musculoskeletal tuberculous infections, neither tuberculous arthritis- nor osteomyelitis-associated. We presented one extremely rare case of primary tuberculous myofasciitis manifesting painful swelling on the left thigh without previous history of tuberculous infection in a patient with dermatomyositis. We also lodged several predisposing symptoms, such as gradually enlarging swelling of extremity within months and proximal muscle painful weakness with prompt response to steroid, which may help early diagnosing of analogous extraspinal musculoskeletal tuberculosis in dermatomyositis patients; and seasonable administrating of anti-TB agents could prevent deterioration in advance.

The effect of the carcinogen safrole on intracellular Ca2+ movement in renal tubular cells has not been explored previously. The present study examined whether safrole could alter Ca2+ handling in Madin-Darby canine kidney (MDCK) cells. Cytosolic free Ca2+ levels ([Ca2+]i) in populations of cells were measured using fura-2 as a fluorescent Ca2+ probe. Safrole at concentrations above 33 microM increased [Ca2+]i in a concentration-dependent manner with an EC50 value of 400 microM. The Ca2+ signal was reduced by 90% by removing extracellular Ca2+, but was not affected by nifedipine, verapamil, or diltiazem. Addition of Ca2+ after safrole had depleted intracellular Ca(2+)-induced dramatic Ca2+ influx, suggesting that safrole caused store-operated Ca2+ entry. In Ca(2+)-free medium, after pretreatment with 650 microM safrole, 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) failed to release more Ca 2+. Inhibition of phospholipase C with 2 microM U73122 did not affect safrole-induced Ca2+ release. Trypan blue exclusion assays revealed that incubation with 650 microM safrole for 30 min did not kill cells, but killed 70% of cells after incubation for 60 min. Collectively, the data suggest that in MDCK cells, safrole induced a [Ca2+] increase by causing Ca2+ release from the endoplasmic reticulum in a phospholipase C-independent fashion, and by inducing Ca2+ influx via store-operated Ca2+ entry. Furthermore, safrole can cause acute toxicity to MDCK cells.

Background The thoracodorsal artery perforator flap was first introduced in 1995. Many authors focused on using anatomical landmarks to identify skin perforators and on thinning procedures for a skin paddle. In this study, we used the superthin free thoracodorsal artery perforator flap for resurfacing shallow defects of the extremities in 10 patients. Methods Two anatomical landmarks previously presented by other authors were used for the guidance of flap elevation. The first landmark represented the site of the proximal skin perforator originating from the descending branch of the thoracodorsal artery, and the second landmark marked the site of the thoracodorsal neurovascular hilum. Ten flaps were performed, and all skin perforators originated from the descending branch of the thoracodorsal artery. The skin paddle carried only skin and the superficial adipose layer, and the largest skin paddle was 23 x 9 cm. Results All defects were resurfaced with smooth contour, except for one flap in which the authors encountered flap tip superficial necrosis. The operative findings did not coincide with the first anatomical landmark in five patients. Likewise, these findings were not consistent with the second landmark in eight patients. In four patients, the thoracodorsal neurovascular hilum was located at the same level of the scapular tip or even above it. These unwanted anatomical variations increased the difficulty of exploration for skin perforators. Conclusions The thoracodorsal artery perforator flap is a feasible choice for shallow defects in superthin form. Using a color Doppler device to identify the skin perforators preoperatively is highly suggested to prevent unexpected anatomical variations.

The effect of the oxidizing agent thimerosal on cytosolic free Ca(2+) concentration ([Ca(2+)]i) and proliferation has not been explored in human osteoblast-like cells. This study examined whether thimerosal alters Ca(2+) levels and causes cell death in MG63 human osteosarcoma cells. [Ca(2+)]i and cell death were measured using the fluorescent dyes fura-2 and WST-1, respectively. Thimerosal at concentrations above 5 microM increased [Ca(2+)]i in a concentration-dependent manner. The Ca(2+) signal was reduced by 80% by removing extracellular Ca(2+). The thimerosal-induced Ca(2+) influx was sensitive to blockade of La(3+), and dithiothreitol (50 microM) but was insensitive to nickel and several L-type Ca(2+) channel blockers. After pretreatment with 1 microM thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor), thimerosal failed to induce [Ca(2+)]i rises. Inhibition of phospholipase C with 2 microM U73122 did not change thimerosal-induced [Ca(2+)]i rises. At concentrations of 5, 10 and 20 microM thimerosal killed 33, 55 and 100% cells, respectively. The cytotoxic effect of 5 microM thimerosal was reversed by 54% by prechelating cytosolic Ca(2+) with BAPTA. Collectively, in MG63 cells, thimerosal induced a [Ca(2+)]i rise by causing Ca(2+) release from endoplasmic reticulum stores and Ca(2+) influx from extracellular space. Furthermore, thimerosal can cause Ca(2+)-related cytotoxicity in a concentration-dependent manner.

Although autogenous tissue can be used to replace unsatisfactory prosthetic breast reconstructions in mastectomy patients, because of the magnitude, complexity, and many potential complications associated with the procedure, combined with a long-term recovery, the use of an implant to replace the mastectomy defect is still the most common method for paraffinoma breast treatment. Between July of 1996 and June of 2003, 21 paraffinoma breast patients underwent bilateral pedicle transverse rectus abdominis myocutaneous (TRAM) flap reconstruction. There were 10 primary cases that had never been treated before this visit, including a case of unilateral associated breast cancer. There were also 11 secondary cases that had prostheses implanted after removal of materials injected in other clinics. The diagnoses included unacceptable breast contour in 11 patients, breast hardening in 11 patients, palpable nodules in five patients, nipple malposition in four patients, prominent scarring in three patients, breast skin necrosis in one patient, and nipple necrosis in one patient. A 100 percent flap survival rate with no clinical fat necrosis was achieved. There were 11 of 21 abdominal hypertrophic scars, six of 21 prechest (anterior surface of the thorax) hypertrophic scars, and no abdominal hernia; the symmetry satisfaction rate was 100 percent among primary cases and nine of 11 in secondary cases. The breast softness satisfaction rate in primary cases was also 100 percent and nine of 11 for secondary cases. Excellent cosmetic results were achieved in all patients (42 breasts in total). The unfavorable results of the secondary cases (patients with previous treatments) indicate that it is impossible to completely remove all of the injected foreign body by resection. This also means that scar appearance can only be minimized if resection of the entire paraffinoma is performed through a periareolar incision. The excellent results of the primary cases show that immediate autogenous tissue reconstruction should be the first alternative and is the best option for treating foreign-body granuloma breast, given that autogenous tissue is available. Similarly, the results of the secondary cases also demonstrate that autogenous tissue reconstruction could be considered in reversing some unfavorable results of past treatments.

The free fibular osteocutaneous flap is often used in the reconstruction of composite oromandibular defects. In contrast, the lateral calcaneal flap has never been used in oromandibular reconstruction. On the basis of their anatomic continuity, the authors combined the free fibular osteocutaneous flap with the lateral calcaneal skin paddle to obtain 2 adjoining flaps in different anatomic areas with the same vascular axis. The authors report their experience in 3 patients with composite oromandibular defects. In 1 patient without an outer skin defect, only a sensory lateral calcaneal skin paddle with a fibular osseus flap was harvested. The lateral calcaneal flap carried the sural nerve as a sensory flap in 2 patients, and the result was passable. Only 1 patient encountered superficial flap tip necrosis at the lateral calcaneal skin paddle, and recovered well after wound care. In conclusion, a flap with many specific features is a feasible choice for reconstruction of composite oromandibular defects.

The use of hypertonic saline (HTS) resuscitation in major trauma patients is still controversial. The objective of this study is to determine if inhibition of inducible nitric oxide synthase (iNOS) to stabilize the endothelial permeability and to retain HTS in the vascular space will reverse its exacerbating effect on burn-induced lung damage. In Experiment 1, specific pathogen-free (SPF) rats underwent 35% total body surface area (TBSA) burn and were resuscitated with 7.5 mL/kg HTS (7.5% NaCl), 7.5 mL/kg saline, or 50 mL/kg saline (nearly equal sodium load as HTS) via femoral veins for 15 min immediately after the burn. In Experiment 2, S-methylisothiourea (SMT) (7.5 mg/kg, i.p.) was given immediately after the burn to rats from the different groups of Experiment 1. At 8 h after the burn, the permeability and myeloperoxidase (MPO) activity of lung tissues were determined, and plasma samples were assayed for peroxynitrite levels. Burn significantly induced lung MPO activity, lung permeability, and blood dihydrorhodamine 123 (DHR 123) oxidation in rats. HTS administration after burn significantly increased the blood DHR 123 oxidation level, lung MPO activity, lung permeability, and inflammatory cell infiltration in comparison with those of burn plus 7.5 mg/kg saline and burn plus 50 mL/kg saline rats. In contrast, burn plus SMT rats with HTS injection showed significant 54%, 11%, and 35% decreases in blood DHR 123 oxidation level, lung MPO activity, and lung permeability, respectively, in comparison with burn plus SMT plus 7.5 mg/kg saline rats. In conclusion, restoration of extracellular fluid in early burn shock with HTS supplementation significantly exacerbates burn-induced lung neutrophil deposition, lung hyperpermeability, and blood peroxynitrite production. Inhibition of iNOS before HTS supplementation reverses the deteriorating effects of HTS on thermal injury-induced lung damage to beneficial ones. Using HTS in thermal injury resuscitation without the inhibition of iNOS is dangerous.

1. The effect of maprotiline, an antidepressant, on human prostate cells is unclear. In the present study, the effect of maprotiline on [Ca 2 + ] i and growth in PC3 human prostate cancer cells was measured using the fluorescent dyes fura-2 and tetrazolium, respectively. 2. Maprotiline caused a rapid, concentration-dependent increase in [Ca 2 + ] i (EC 5 0 = 200 μmol/L). The maprotiline-induced [Ca 2 + ] i increase was reduced by removal of extracellular Ca 2 + or pretreatment with nicardipine. 3. The maprotiline-induced Mn 2 + influx-associated fura-2 fluorescence quench directly suggests that maprotiline caused Ca 2 + influx. 4. In Ca 2 + -free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2 + -ATPase, caused a monophasic [Ca 2 + ] i increase, after which the effects of maprotiline of increasing [Ca 2 + ] i were abolished. In addition, pretreatment with maprotiline reduced a major portion of the thapsigargin-induced increase in [Ca 2 + ] i . 5. U73122, an inhibitor of phospholipase C, abolished the ATP (but not maprotiline)-induced increase in [Ca 2 + ] i . 6. Overnight incubation with 1-10 μmol/L maprotiline did not alter cell proliferation, although incubation with 30-50 μmol/L maprotiline decreased cell proliferation. 7, These findings suggest that maprotiline rapidly increases [Ca 2 + ] i in human prostate cancer cells by stimulating both extracellular Ca 2 + influx and intracellular Ca 2 + release and that it may modulate cell proliferation in a concentration-dependent manner.

In Madin-Darby canine kidney (MDCK) cells, the effect of 2-O-methyl PAF, an inactive analogue of platelet activating factor (PAF), on intracellular Ca 2+ concentration ([Ca 2+ ] i ) was measured by using the Ca 2+ -sensitive fluorescent dye fura-2. 2-O-methyl PAF (≥15 μM) caused a rapid rise of [Ca 2+ ] i in a concentration-dependent manner. 2-O-methyl PAF-induced [Ca 2+ ] i rise was partly reduced by removal of extracellular Ca 2+ . 2-O-methyl PAF-induced extracellular Ca 2+ influx was also suggested by Mn 2+ influx-induced fura-2 fluorescence quench. The 2-O-methyl PAF-induced Ca 2+ influx was blocked by nifedipine, verapamil and diltiazem. In Ca 2+ -free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2+ -ATPase, caused a monophasic [Ca 2+ ] i rise, after which 2-O-methyl PAF failed to increase [Ca 2+ ] i ; also, pretreatment with 2-O-methyl PAF depleted thapsigargin-sensitive Ca 2+ stores. U73122, an inhibitor of phospholipase C, abolished ATP (but not 2-O-methyl PAF)-induced [Ca 2+ ] i rise. These findings suggest that 2-O-methyl PAF evokes a rapid increase in [Ca 2+ ] i in renal tubular cells by stimulating both extracellular Ca 2+ influx and intracellular Ca 2+ release.

The differences in the intracellular Ca(2+) responses to hormones in platelets from systemic lupus erythematosus (SLE) patients compared to normal humans have not been explored. This study examined the Ca(2+) signaling and density of platelets in normal, inactive and active SLE patients. The platelet number per mul in inactive and normal groups did not differ, whereas the number in active SLE patients was smaller than the other two groups by 60%. The intracellular free Ca(2+) levels ([Ca(2+)](i)) in response to stimulation of four endogenous Ca(2+) mobilizing hormones, 100 microM arachidonic acid (AA), 10 microM ADP, 10 nM platelet activation factor (PAF) and 1 microM thrombin, were investigated using the Ca(2+)-sensitive fluorescent dye, fura-2. The AA-induced [Ca(2+)](i) rises in normal and inactive groups were similar. In contrast, the AA-induced [Ca(2+)](i) rises in the active SLE group were significantly smaller than in the normal and inactive groups. The defect in the AA-induced [Ca(2+)](i) rises in active SLE groups appears to be caused by defective Ca(2+) influx and Ca(2+) releasing pathways because the AA-induced responses were not altered by removal of extracellular Ca(2+), whereas the AA-induced responses in normal and inactive SLE groups were reduced by removal of extracellular Ca(2+), and the AA-induced Ca(2+) release was smaller in the active SLE group. PAF, ADP and thrombin all induced [Ca(2+)](i) rises in the three groups, but no significant differences were found among the three groups. Together, the results indicate that cell density and Ca(2+) signaling in platelets from active SLE patients are altered in response to particular stimulators. In these regards, platelets from inactive SLE patients appear to be similar to those from normal humans.

In human breast cancer cells, the effect of the widely prescribed estrogen diethylstilbestrol (DES) on intracellular Ca2+ concentrations ([Ca2+]i) and cell viability was explored by using fura-2 and trypan blue exclusion, respectively. DES caused a rise in [Ca2+]i in a concentration-dependent manner (EC50 = 15 microM). DES-induced [Ca2+]i rise was reduced by 80 % by removal of extracellular Ca2+. DES-induced Mn(2+)-associated quench of intracellular fura-2 fluorescence also suggests that DES induced extracellular Ca2+ influx. In Ca(2+)-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca(2+)-ATPase, caused a monophasic [Ca2+]i rise, after which the increasing effect of DES on [Ca2+]i was greatly inhibited. Conversely, pretreatment with DES to deplete intracellular Ca2+ stores totally prevented thapsigargin from releasing more Ca2+, whereas ionomycin added afterward still released some Ca2+. These findings suggest that in human breast cancer cells, DES increases [Ca2+]i by stimulating extracellular Ca2+ influx and also by causing intracellular Ca2+ release from the endoplasmic reticulum. Acute trypan blue exclusion studies suggest that 10-20 NM DES killed cells in a time-dependent manner.

In Madin-Darby canine kidney (MDCK) cells, the effect of maprotiline, an antidepressant, on intracellular Ca(2+) concentration ([Ca(2+)](i)) was measured using fura-2. Maprotiline (2.5 microM) caused a rapid rise of [Ca(2+)](i) in a concentration-dependent manner (EC(50) 200 microM). Maprotiline-induced [Ca(2+)](i) rise was reduced by removal of extracellular Ca(2+) or by addition of La(3+), but was not altered by voltage-gated Ca(2+)-channel blockers. Maprotiline-induced Mn(2+) influx-associated fura-2 fluorescence quench directly suggests that maprotiline caused Ca(2+) influx. In Ca(2+)-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca(2+)-ATPase, caused a monophasic [Ca(2+)](i) rise, after which the increasing effect of maprotiline on [Ca(2+)](i) was nearly abolished; also, pretreatment with maprotiline reduced a portion of thapsigargin-induced [Ca(2+)](i) rise. U73122, an inhibitor of phospholipase C, abolished [Ca(2+)](i) rise induced by ATP (but not by maprotiline). Overnight incubation with 1-10 microM maprotiline enhanced cell viability, but 20-50 microM maprotiline decreased it. These findings suggest that maprotiline rapidly increases [Ca(2+)](i) in renal tubular cells by stimulating both extracellular Ca(2+) influx and intracellular Ca(2+) release, and may modulate cell proliferation in a concentration-dependent manner.

The effect of nordihydroguaiaretic acid (NDGA), a compound commonly used as a lipoxygenases inhibitor, on intracellular free Ca2+ levels ([Ca2+]i) in PC3 human prostate cancer cells was investigated. [Ca2+]i was measured by using the Ca2+ -sensitive dye fura-2. NDGA increased [Ca2+]i in a concentration-dependent manner with an EC50 of 30 microM. The Ca2+ signal comprised a gradual and sustained increase. Removal of extracellular Ca2+ partly decreased the NDGA-induced [Ca2+]i increase, suggesting that the Ca2+ signal was due to both extracellular Ca2+ influx and intracellular Ca2+ release. NDGA-induced Ca2+ influx was independently confirmed by measuring NDGA-induced Mn2+ -coupled quench of fura-2 fluorescence. The NDGA-induced Ca2+ influx was not affected by L-type Ca2+ channel blockers. In Ca2+ -free medium, the NDGA-induced [Ca2+]i increase was abolished by pretreatment with 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor), and conversely, pretreatment with NDGA abolished thapsigargin-induced [Ca2+]i increase. NDGA-induced intracellular Ca2+ release was not altered by inhibition of phospholipase C. Overnight treatment with 20-50 microM NDGA inhibited cell proliferation rate in a concentration-dependent manner. Several other lipoxygenases inhibitors did not alter [Ca2+]i. Collectively, this study shows that in prostate cells, NDGA induced a [Ca2+]i increase via releasing stored Ca2+ from the endoplasmic reticulum in a manner independent of phospholipase C activity, and by causing Ca2+ influx. NDGA also caused cytotoxicity at higher concentrations.

This study was conducted to demonstrate the burn-induced lung neutrophil deposition and damage in rats is affected by the nitric oxide (NO)-dependent downstream cGMP signaling. In experiment 1, 1H-[1,2,4] oxadiazolo [4,3-alpha] quinoxalin-1-one (ODQ) was given (20 mg/kg i.p.) to specific pathogen-free Sprague-Dawley rats immediately postburn to suppress the guanylate cyclase (GC) activity. At 8 h after burn, blood was assayed for the peroxynitrite-mediated dihydrorhodamine 123 (DHR 123) oxidation and lung tissues were harvested for myeloperoxidase (MPO) determination and histological studies. Pulmonary microvascular dysfunction was quantified by measuring the extravasations of Evans blue dye. In experiment 2, Sodium nitroprusside (SNP) was given (2 mM, i.p.) to elevate cGMP levels and ODQ (20 mg/kg, i.p.) or methylene blue (100 microM, i.p.) or saline was given. The animals were sacrificed 4 h after injection and lung tissues were harvested for iNOS mRNA study. The MPO activity in lung, blood DHR 123 oxidation level, and lung permeability increased up to 2-fold, 4-fold, and 2.5-fold after burn. Inhibition of GC by ODQ administration significantly decreased MPO activity, blood DHR 123 oxidation, and lung permeability by 55%, 66%, and 53%, respectively, and markedly decreased the thermal injury-induced perivascular and interstitial inflammatory cell infiltration and septum edema. The protective effects of ODQ were comparable to the use of selective iNOS inhibitor as demonstrated previously. Furthermore, ODQ decreased the burn or SNP-induced iNOS mRNA levels at 4 h after burn. These findings suggest that burn-induced lung dysfunction is mediated by the NO/cGMP system because it is abolished by application of either iNOS inhibitor or GC inhibitor. Also, the beneficial effect of ODQ is partly due to the attenuation of burn-induced iNOS expression by GC inhibition.

Intestinal ischemia and reperfusion injury was known to cause postinjury multiple organ failure by neutrophil and unclear nonneutrophil factors. Peroxynitrite formed by the rapid reaction between superoxide and nitric oxide, is a toxic substance that contributes to tissue injury in a number of biological systems. In this study, the role of nitric oxide and neutrophils on lung damage after burn was investigated.Prospective, experimental study.Research laboratory at a university hospital.Thermal injury models in the rat.In experiment 1, specific pathogen-free Sprague-Dawley rats underwent 35% total body surface area burn. At 4, 8, 16, and 24 hrs after burn, intestinal mucosa and lung tissue were harvested for myeloperoxidase (MPO) assay, blood was collected for measurement of peroxynitrite-mediated oxidation of dihydrorhodamine 123, and pulmonary microvascular dysfunction was quantified by measuring the extravasation of Evans blue dye. In experiment 2, polymorphonuclear granulocyte antibody (0.12 mL/100 g administered intraperitoneally 16 hrs before burn), S-methylisothiourea (7.5 mg/kg, intraperitoneally, immediately after burn), a specific inducible nitric oxide synthase inhibitor, and sterile saline (15 mL/kg, intraperitoneally, immediately after burn) were given to different groups of thermally injured animals individually. The plasma dihydrorhodamine 123 oxidation level, intestinal and lung MPO activity, lung permeability, and lung histology were evaluated at 8 hrs after burn. The cellular localization of nitrotyrosine, a marker for peroxynitrite reactivity, was also examined by immunostaining. In experiment 3, 3-morpholinosydnonimine (10 mM, intraperitoneally), a peroxynitrite donor, was given to nonburned rats to examine the peroxynitrite effect on lung inducible nitric oxide synthase expression.The level of MPO activity in intestine and lung, blood dihydrorhodamine 123 oxidation, and lung permeability were increased up to 2-fold, 2.5-fold, 2-fold, and 2-fold of normal, respectively, at 8 hrs after burn. S-methylisothiourea injection significantly decreased (p.05) 31% of the lung MPO activity, 41% of the blood peroxynitrite level, 54% of the lung permeability, and the lung peroxynitrite production in burned rats. Polymorphonuclear granulocyte antibody pretreatment significantly decreased 60% of the intestinal MPO, 92% of the blood peroxynitrite level, and 56% the lung MPO activity in burned rats, but the lung permeability was only slightly decreased by polymorphonuclear granulocyte antibody pretreatment. Furthermore, 3-morpholinosydnonimine increased the lung inducible nitric oxide synthase messenger RNA levels.Thermal injury induces blood dihydrorhodamine 123 oxidation, intestinal and lung neutrophil deposition, lung nitrotyrosine production, and lung damage. Both specific inhibition of inducible nitric oxide synthase and polymorphonuclear granulocyte antibody pretreatment decrease blood dihydrorhodamine 123 oxidation and intestinal and lung neutrophil deposition, but only inducible nitric oxide synthase inhibition with S-methylisothiourea reduces lung peroxynitrite production and thermal injury-induced lung damage. Nitric oxide and the ensuing peroxynitrite production in lung play a more important role than neutrophil in contributing to thermal injury-induced lung damage.

The double-paddle peroneal tissue transfer is a useful technique for reconstructing the extensive and complex defect that results after ablative surgery for oral cancer. It can facilitate the design and inset of the skin paddle and avoid the need for a second free flap. The two skin paddles can be based on either two cutaneous perforators of the peroneal vessels or two branches of a single cutaneous perforator. The authors report their experience with double-paddle peroneal tissue flaps (10 fasciocutaneous and five osteocutaneous) in 15 patients. The largest double paddle used was (16 X 9) (15 X 6) cm, and the smallest one was (7 X 5.5) (4.5 X 4) cm. All flaps were used for both intraoral and extraoral defect reconstruction. There was one single skin paddle necrosis caused by erroneous manipulation of the flap 1 week after the operation; however, the skin paddle had survived completely before the manipulation. All other flaps survived completely, with a good to excellent appearance, and no patient had a significant gait disturbance after the operation.

Calcium (Ca 2+ ) influx into human polymorphonuclear cells (PMNs) in response to N-formyl-Met-Leu-Phe (fMLP) and platelet-activating factor (PAF) stimulation was studied. Whole blood was taken by venous puncture from healthy human volunteers. PMNs were isolated, diluted, and incubated with 2 μM fura-2 AM. The cytosolic free calcium concentration, [Ca 2+ ] i , in human neutrophils was determined by microfluorometry. We found that the net area under the fMLP- or PAF-induced [Ca 2+ ] i rise curve in Ca 2+ -free medium decreased to 75% or 30% of the area under the curve in Ca 2+ medium. Treatment of PMNs with phorbol myristate acetate (PMA), a protein kinase C activator, completely abolished the intracellular Ca 2+ level stimulated by PAF, but not the intracellular Ca 2+ level stimulated by fMLP. Treatment of PMNs with PAF did not abolish the intracellular Ca 2+ level elevation stimulated by fMLP. In addition, treatment of PMNs with fMLP did not abolish intracellular Ca 2+ level elevation stimulated by PAF. Loperamide, a positive modulator for store-operated calcium (SOC) channels, elicited an increase in intracellular calcium after the activation of SOC channels stimulated by fMLP or PAF. After the addition of guanosine 3',5'-cyclic monophosphate, N 2 ,2'-O-Dibutyryl-, sodium salt (db-cGMP), the initial increase of PAF- or fMLP-induced PMNs intracellular Ca 2+ fluorescences was well preserved, but the slope and the peak height of fluorescence curves declined compared with the curves without db-cGMP. In conclusion, we found that PAF and fMLP regulate the Ca 2+ influx of PMNs in different ways. Most of the PAF-induced [Ca 2+ ] i rise resulted from Ca 2+ influx, and most of the fMLP-induced [Ca 2+ ] i elevation resulted from intracellular stores release. The initial mobilization of intracellular Ca 2+ stores in PAF-stimulated signals is mediated by protein kinase C (PKC) phosphorylation, but not in fMLP-stimulated route. SOC channels are present and important in the fMLP- or PAF-induced PMNs Ca 2+ influx. There was no apparent cross-regulation between PAF- and fMLP-stimulated intracellular Ca 2+ influx.

Background The aim of this study was to evaluate the recurrence pattern after skin-sparing mastectomy (SSM) and immediate breast reconstruction (IBR) using transverse rectus abdominis musculocutaneous (TRAM) flap in patients with invasive breast cancer. Methods From 1995 to 2010, patients with invasive breast cancer who underwent SSM followed by IBR using TRAM flap were retrospectively reviewed. The pattern of the first recurrence event was recorded. Results We identified 249 consecutive patients with invasive breast cancer, two-thirds of whom (67.1%) were diagnosed with stage II or stage III disease. During a median follow-up period of 53 months, three (1.2%) local, 13 (5.2%) regional, 34 (13.7%) distant, and five (2.0%) concurrent locoregional and distant recurrences were observed. The median time to recurrences was 26 months (range, 2 to 70 months) for all recurrences, 23 months (range, 2 to 64 months) for locoregional recurrences, and 26 months (range, 8 to 70 months) for distant recurrences. All local recurrent lesions were detectable by careful physical examination, and detection of local recurrence suggested the presence of distant metastasis (60.0%). In contrast to distant metastasis, the risk of locoregional recurrence did not increase significantly with an increase in disease stage. The 5-year overall, locoregional relapse-free, and distant relapse-free survival rates were 89.7%, 90.8%, and 81.6%, respectively. Conclusions SSM followed by immediate reconstruction using TRAM flap is an oncologically safe procedure even in patients with advanced-stage disease. Detection of local recurrence is crucial and can be aided by a thorough physical examination.

e15562 The optimal management of LAPC remains controversial. The efficacy of CCRT is hampered by early systemic dissemination. To treat undetectable metastases and to select patients who are likely to benefit from CCRT, ICT followed by CCRT has recently been extensively evaluated in LAPC. Recently, we showed that triplet chemotherapy consisting of gemcitabine 800 mg/m2 (10 mg/m2/min) followed by oxaliplatin 85 mg/m2 and 48-hour infusion of 5-FU/LV 3,000 and 150 mg/m2 Q 2 weeks, the GOFL regimen, is feasible and active for pts with APC. Chemo-naïve pts with histo- /cytologically proven unresectable LAPC, and bi-dimensionally measurable diseases were eligible. Patients who did not experience disease progression (PD) after 6 cycles of GOFL would had CCRT consisting of weekly gemcitabine 400mg/m2 plus 50.4Gy/28 fractions of radiation 4–6 weeks later. After CCRT, pts were re-evaluated for surgical intervention and those with unresectable disease would continue GOFL until PD, unacceptable toxicity, patient's refusal or death. Among the 50 enrolled pts (24F/26M, median age 58.5 years), 48 had definitively unresectable diseases. After induction GOFL, 16 (32%) pts were off-studied because of PD in 12 (24%) and ICT-related toxicity in 4 (8%, PD/UE group). Among the 34 (68%) with objective response or stable disease after 6 cycles of ICT, 27 (54%) who completed the assigned multimodality treatment are categorized as CCRT group; whiles 7 (14%) who either declined CCRT (in 5) or still on ICT (in 2) are categorized as non-CCRT group. The median PFS and OS for the ITT population were 9.1 and 14.5 months, respectively. The median PFS for PD/UE, non-CCRT and CCRT groups were 2.1, 8.2 and 12.8 months, respectively; whiles the OS were 8.5, 15.0 and 18.3 months, respectively. The most common grade 3–4 toxicities were neutropenia (40%), anemia (14%), infection (16%), nausea (26%), vomiting (20%). Grade 2–3 peripheral neuropathy was observed in 7 of 29 who received post-CCRT GOFL. Conclusion: Three months of triplet ICT followed by gemcitabine-based CCRT is feasible and likely to prolong the survival of selected LAPC pts. Prospective, randomization study is warranted to validate the findings. No significant financial relationships to disclose.

The use of albumin in the early resuscitation formula after major burn has been forbidden because of its damaging effect on the gut barrier function. We hypothesize that inhibition of the inducible isoform of nitric oxide synthase to stabilize endothelial permeability and to retain albumin in the vascular space will ameliorate the major trauma-induced gut barrier dysfunction.In experiment 1, specific pathogen-free rats undergoing 35% total body surface area burn or sham burn were given equal volumes (7.5 mL/kg) of isotonic sodium chloride solution or albumin from femoral veins for fluid resuscitation at 0, 4, or 8 hours after burn. In experiment 2, intraperitoneal S-methylisothiourea sulfate (7.5 mg/kg) was given immediately after burn to rats from different groups, as in experiment 1 (SMT groups). At 24 hours after burn, the intestinal mucosa was assayed for myeloperoxidase activity as an index for neutrophil sequestration, the distribution of fluorescein isothiocyanate-dextran across the lumen of small intestine was determined to evaluate the intestinal permeability, and bacterial translocation (BT) to the mesenteric lymph nodes (MLNs) and histological findings in the ileum were also examined.Compared with sham burn, burn induced significant increases in intestinal mucosa myeloperoxidase activity, intestinal permeability, BT to the MLNs, and villi sloughing in rats. Albumin administration at 0 or 4 hours after burn enhanced the increases in neutrophil sequestration, permeability, and villi sloughing compared with saline injection at the same times. In contrast, injection of albumin in the burn-SMT group did not aggravate these changes in intestinal myeloperoxidase activity, intestinal permeability, BT to the MLNs, and villi edema. Burn-SMT rats with albumin injections at 4 or 8 hours after burn showed significant 35% and 52% decreases, respectively, in intestinal permeability compared with burn-SMT-saline rats. Use of albumin at 8 hours after burn in combination with S-methylisothiourea significantly attenuated BT to the MLNs and reduced villi edema.Early albumin resuscitation aggravated the burn-induced gut damage. Albumin administration and inhibition of the inducible isoform of nitric oxide synthase in combination decreased burn-induced gut barrier dysfunction and reversed the damaging effect of albumin on gut barrier function and decreased BT.

The effectiveness of a modified glucose-insulin-potassium (GIK) formula, which was derived from the results of a previous study (the maximal glucose disposal rate of 400 mg/M2/min required an insulin infusion rate at 1,200 mU/M2/min and KCl supplement rate at 0.08 mEq/M2/min), was evaluated in the treatment of extensive acute burn injury (EABI) in dogs. Under anesthesia initially with intravenous sodium pentobarbital 35 mg/kg followed by a maintenance dose of 5 mg/kg/hr, a third-degree burn of about 50% of the total body surface was created by acetylene torch over the ventral wall of the chest and abdomen. Cardiovascular parameters including heart rate, mean arterial blood pressure, pulmonary wedge pressure, cardiac index, and cardiac contractility (dP/dt of left ventricular pressure), as well as blood chemical data of pH value and K+ concentration were monitored. The present GIK therapy in EABI dogs effectively prevented a decrease in cardiac function, markedly enhanced cardiac function, steadily prolonged cardiac enhancement, and safely avoided hypoglycemic attack.