Cystic pancreatic neuroendocrine tumors: endoscopic ultrasound and fine-needle aspiration characteristics.

BACKGROUND AND STUDY AIMS: Limited data are available on the endoscopic ultrasound (EUS) and fine-needle aspiration (FNA) characteristics of cystic pancreatic neuroendocrine tumors (CPanNets). The aims of this study were to describe the EUS and FNA characteristics of pathologically confirmed CPanNets and to compare these characteristics with mucinous cysts from matched patients. PATIENTS AND METHODS: From an EUS - FNA database (between 1999 and 2011), 19 patients with a pathologically confirmed CPanNet were identified. Patient characteristics, cyst fluid carcinoembryonic antigen (CEA) levels, pathology, and EUS findings were analyzed. For comparison, age- and sex-matched patients with mucinous cysts were randomly chosen from the same database. RESULTS: Of the 19 patients, two had multiple endocrine neoplasia type 1 and two had metastases. The median diameter of the lesions was 24 mm. EUS revealed unilocular lesions in 7 patients, thinly septated lesions with thin walls in 1, and mixed solid-cystic lesions in 11. EUS - FNA cytology confirmed neoplasm in 12 of the 19 patients (63.2 %). The median cyst fluid CEA level (n = 15) was 1.1 ng/mL (range 0.3 - 500 ng/mL). Compared with matched patients with mucinous cysts, the median cyst fluid CEA was lower (1.1 ng/mL vs. 400 ng/mL), thick walls were more common (66.7 % vs. 13.3 %), and diagnostic cytology was more likely (73.3 % vs. 20.0 %). CONCLUSIONS: Analysis of EUS and FNA results showed that the cyst fluid from CPanNets had a lower CEA concentration, a higher frequency of thick walls on EUS, and higher diagnostic cytology compared with mucinous cysts. These findings may aid in the diagnosis of CPanNets.

Time-lapse, single cell based confocal imaging analysis of caspase activation and phosphatidylserine flipping during cellular apoptosis.

Apoptosis is an important phenomenon for investigating the efficacy of anti-cancer drug candidates. The conventional assays for cellular apoptosis, such as enzyme-linked immunosorbent assay, absorbance monitoring for the activity of caspase, and flow cytometric assay, have focused only on biochemical events. We investigated the staurosporine (STS)-induced apoptosis of the murine macrophage RAW-264.7 cell using a cell based bioimaging technique. Using time-lapse confocal microscopy, we monitored caspase-3 activation during apoptosis by imaging the translocation of green fluorescent protein from the cytosol to the nuclei. Five hours after 1 µM STS treatment, caspase-3 was observed to be activated and membrane blebbing was observed simultaneously. Also, the loss of phosphatidylserine (PS) asymmetry in the phospholipid bilayer of plasma membrane during early apoptosis was monitored by imaging annexin-V labeled with fluorescein isocyanate binding to the externalized PS at various concentrations of STS. Moreover, disintegration of the plasma membrane during late apoptosis was confirmed using a nuclear dye, propidium iodide. The single cell based bioimaging data agreed well with those of the biochemical assays for caspase activation and morphological observation for membrane integrity.

Artemisia fukudo essential oil attenuates LPS-induced inflammation by suppressing NF-kappaB and MAPK activation in RAW 264.7 macrophages.

In the present study, the chemical constituents of Artemisia fukudo essential oil (AFE) were investigated using GC-MS. The major constituents were alpha-thujone (48.28%), beta-thujone (12.69%), camphor (6.95%) and caryophyllene (6.01%). We also examined the effects of AFE on the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and IL-6, in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Western blotting and RT-PCR tests indicated that AFE has potent dose-dependent inhibitory effects on pro-inflammatory cytokines and mediators. We investigated the mechanism by which AFE inhibits NO and PGE(2) by examining the level of nuclear factor-kappaB (NF-kappaB) activation within the mitogen-activated protein kinase (MAPK) pathway, which is an inflammation-induced signal pathway in RAW 264.7 cells. AFE inhibited LPS-induced ERK, JNK, and p38 phosphorylation. Furthermore, AFE inhibited the LPS-induced phosphorylation and degradation of Ikappa-B-alpha, which is required for the nuclear translocations of the p50 and p65 NF-kappaB subunits in RAW 264.7 cells. Our results suggest that AFE might exert an anti-inflammatory effect by inhibiting the expression of pro-inflammatory cytokines. Such an effect is mediated by a blocking of NF-kappaB activation which consequently inhibits the generation of inflammatory mediators in RAW264.7 cells. AFE may be useful for treating inflammatory diseases.

Branch duct intraductal papillary mucinous neoplasms in a retrospective series of 190 patients.

BACKGROUND: A consensus conference has recommended close observation of branch duct intraductal papillary mucinous neoplasms (BD-IPMNs) smaller than 30 mm, without symptoms or mural nodules. This study investigated whether these recommendations could be validated in a single-centre experience of BD-IPMNs. METHODS: Some 190 patients with radiological imaging or histological findings consistent with BD-IPMN were enrolled between 1998 and 2005. Those with less than 6 months' follow-up and no histological confirmation were excluded. RESULTS: BD-IPMN was diagnosed by computed tomography and pancreatography in 105 patients and pathologically in 85. Eighteen patients had adenoma, 53 borderline malignancy, five carcinoma in situ and nine invasive carcinoma. Findings associated with malignancy were the presence of radiologically suspicious features (P < 0.001) and a cyst size of at least 30 mm (P = 0.001). Had consensus guidelines been applied, 54 patients would have undergone pancreatic resection, whereas only 28 of these patients actually had a resection; 12 of the latter patients had a malignancy compared with none of the 26 patients who were treated conservatively. CONCLUSION: A simple increase in cyst size is not a reliable predictor of malignancy. Observation is recommended for patients with a BD-IPMN smaller than 30 mm showing no suspicious features on imaging.

Efficacy of alphas1-casein hydrolysate on stress-related symptoms in women.

OBJECTIVE: To examine the effects of alpha (s1)-casein hydrolysate on females with stress-related symptoms. DESIGN: Double-blind, randomized, crossover, placebo-controlled trial. SETTING: The alpha (s1)-casein hydrolysate was manufactured by INGREDIA (Arras, France) and the placebo was manufactured by DIETAROMA (Bourg, France). Study was designed and performed at PROCLAIM (Rennes, France), and the statistical analyses were performed by D Desor (Nancy, France). SUBJECTS: A total of 63 female volunteers suffering from at least one disorder that may be related to stress such as anxiety, sleep problems and general fatigue. INTERVENTIONS: A total of 63 volunteers participated in a double-blind, randomized, crossover, placebo-controlled study. Subjects were randomly allocated to receive either tablets containing alpha (s1)-casein hydrolysate or placebo at the dose of 150 mg/day for 30 days. After a 3 weeks washout period, they were crossed over for a new 30-day period of tablets intake. The outcome measure was a questionnaire including 44 items of symptoms that may be related stress in which the severity of each sign was evaluated using a 10-degree scale. These measures were studied repeatedly at the day of 0, 15 and 30 after the start of each interventional period. RESULTS: The 30-day treatment by alpha (s1)-casein hydrolysate in females with stress-related symptoms reduced their symptoms, particularly in digestion (P<0.01), cardiovascular (P<0.05), intellectual (P<0.01), emotional (P<0.05) and social problems (P<0.05). CONCLUSION: This study showed that a 30-day ingestion of alpha (s1)-casein hydrolysate decreased the stress-related symptoms in females suggesting that this product may be used as an effective functional ingredient alleviating such symptoms. SPONSORSHIP: This study was partially supported by the INGREDIA of France and Neurobiology Research Program from the Korea Ministry of Science and Technology (2004-01757) of Korea.

Blockade of ionotropic glutamate receptors produces neuronal apoptosis through the Bax-cytochrome C-caspase pathway: the causative role of Ca2+ deficiency.

Blockade of ionotropic glutamate receptors induces neuronal cell apoptosis. We investigated if mitochondria-mediated death signals would contribute to neuronal apoptosis following administration of glutamate antagonists. The administration of MK-801 and CNQX (MK-801/CNQX), the selective antagonists of N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors, produced widespread neuronal death in neonatal rat brain and cortical cell cultures. MK-801/CNQX-induced neuronal apoptosis was prevented by zVAD-fmk, a broad inhibitor of caspases, but insensitive to inhibitors of calpain or cathepsin D. Activation of caspase-3 was observed within 6-12 h and sustained over 36 h after exposure to MK-801/CNQX, which cleaved PHF-1 tau, the substrate for caspase-3. Activation of caspase-3 was blocked by high K+ and mimicked by BAPTA-AM, a selective Ca2+ chelator. Reducing extracellular Ca2+, but not Na+, activated caspase-3, suggesting an essential role of Ca2+ deficiency in MK-801/CNQX-induced activation of caspases. Cortical neurons treated with MK-801/CNQX triggered activation of caspase-9, release of cytochrome c from mitochondria, and translocation of Bax into mitochondria. The present study suggests that blockade of ionotropic glutamate receptors causes caspase-3-mediated neuronal apoptosis due to Ca2+ deficiency that is coupled to the sequential mitochondrial death pathway.

1,2-bis(2-Aminophenoxy)ethane-N,N,N',N'-tetraacetic acid induces caspase-mediated apoptosis and reactive oxygen species-mediated necrosis in cultured cortical neurons.

Sustained alteration in [Ca(2+)]i triggers neuronal death. We examined morphological and signaling events of Ca(2+)-deficiency-induced neuronal death. Cortical cell cultures exposed to 20 microM 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM), an intracellular calcium chelator, underwent neuronal apoptosis within 12 h that was evident by shriveled cell bodies, aggregated and condensed nuclear chromatin, and disrupted nuclear membrane. Thereafter, surviving neurons revealed typical necrosis, accompanied by swelling of cell body and mitochondria, over 24 h. Both apoptosis and necrosis were prevented by inclusion of 1 microg/mL cycloheximide, a protein synthesis inhibitor. Treatment with BAPTA-AM induced translocation of Bax into mitochondria within 4 h and release of cytochrome c from mitochondria over 4-12 h. An active fragment of caspase-3, a downstream mediator of cytochrome c, was observed within 8 h and cleaved PHF-1-positive tau. Administration of zVAD-fmk, a broad inhibitor of caspases, or DEVD-amc, a selective inhibitor of caspase-3, selectively prevented the apoptosis component of BAPTA-AM neurotoxicity. In contrast, BAPTA-AM-induced necrosis was propagated through sequential production of superoxide, mitochondrial and cytoplasmic reactive oxygen species. Combined treatment with caspase inhibitors and antioxidants blocked BAPTA-AM neurotoxicity. The present study suggests that neurons deficient in [Ca(2+)]i undergo caspase-3-mediated apoptosis and reactive oxygen species (ROS)-mediated necrosis.

Proapoptotic effects of tau cleavage product generated by caspase-3.

Using an in vitro translation assay to screen a human brain cDNA library, we isolated the microtubule-associated protein Tau and determined it to be a caspase-3 substrate whose C-terminal cleavage occurred during neuronal apoptosis. DeltaTau, the 50-kDa cleavage product, was detected by Western blot in apoptotic cortical cells probed with anti-PHF-1 and anti-Tau-5 antibodies, but not anti-T-46 antibody which recognizes the C-terminus. Overexpression of DeltaTau in SK-N-BE2(C) cells significantly increased the incidence of cell death. Staurosporine-induced Tau cleavage was blocked by 20 microM z-Asp-Glu-Val-Asp-chloromethylketone, a caspase-3 inhibitor, and in vitro, Tau was selectively cleaved by caspase-3 or calpain, a calcium-activated protease, but not by caspases-1, -8, or -9. (D421E)-Tau, a mutant in which Asp421 was replaced with a Glu, was resistant to cleavage by caspase-3 and tended to suppress staurosporine-induced cell death more efficiently than did wild-type Tau in both transient and stable expression systems. Finally, the incidence of DeltaTau-induced cell death was augmented by expression of Abeta precursor protein (APP) or Swedish APP mutant. Taken together, these results suggest that the caspase-3 cleavage product of Tau may contribute to the progression of neuronal cell death in Alzheimer's disease.

Synergetic activation of p38 mitogen-activated protein kinase and caspase-3-like proteases for execution of calyculin A-induced apoptosis but not N-methyl-d-aspartate-induced necrosis in mouse cortical neurons.

We examined the possibility that p38 mitogen-activated protein kinase and caspase-3 would be activated for execution of apoptosis and excitotoxicity, the two major types of neuronal death underlying hypoxicischemic and neurodegenerative diseases. Mouse cortical cell cultures underwent widespread neuronal apoptosis 24 h following exposure to 10-30 nM calyculin A, a selective inhibitor of Ser/Thr phosphatase I and IIA. Activity of p38 was increased 2-4 h following exposure to 30 nM calyculin A. Addition of 3-10 microM PD169316, a selective p38 inhibitor, partially attenuated calyculin A neurotoxicity. Activity of caspase-3-like proteases was increased in cortical cell cultures exposed to 30 nM calyculin A for 8-16 h as shown by cleavage of DEVD-p-nitroanilide and phosphorylated tau. Proteolysis of tau was completely blocked by addition of 100 microM N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (z-VAD-fmk), a broad-spectrum inhibitor of caspases, but incompletely by 10 microM PD169316. Calyculin A neurotoxicity was partially sensitive to 100 microM z-VAD-fmk. Cotreatment with 10 microM PD169316 and 100 microM z-VAD-fmk showed additive neuroprotection against calyculin A. Neither PD169316 nor z-VAD-fmk showed a beneficial effect against excitotoxic neuronal necrosis induced by exposure to 20 microM NMDA. Thus, caspase-3-like proteases and p38 likely contribute to calyculin A-induced neuronal apoptosis but not NMDA-induced neuronal necrosis.

Submuscular periareolar approach to augmentation mammoplasty in Korean women.

Augmentation mammoplasty can be approached by various methods according to the type of implant and implantation site depending on the status of the patient or surgeon's preference. The advantage for submuscular placement is based on problems associated with subglandular placement, especially capsular contracture and sensory changes in the nipple, and interference with the interpretation of mammograms is avoided. There are fewer complications such as hematoma, infection, and extrusion of the implant with submuscular dissection and relatively avascular, minimal sensory changes in the nipple compared with subglandular approach. The submuscular periareolar approach to augmentation mammoplasty was first described in the 1970s. This approach provides easy access to both the subglandular and subpectoral planes. It also provides a central point of access for creation of the implant pocket, which allows for easier and more accurate dissection in all diameters. The resultant periareolar scar is usually minimal with less injury to breast parenchyme and eventual biopsy or mastectomy incision to be performed through or around the areola. During the period of March 1999 to January 2000, 19 cases of who received submuscular periareolar augmentation mammoplasty under general anesthesia resulted in favorable scars with accurate access to pocket margin, easier dissection, and less bleeding compared with submuscular transaxillary augmentation mammoplasty. In our experience with the submuscular periareolar approach to breast augmentation it was highly versatile, safe, and less painful; postoperative hematoma incidence was greatly reduced and breast tissue injury was minimized.

Molecular cloning of a novel GTP-binding protein induced in fish cells by rhabdovirus infection.

We have cloned and sequenced a cDNA encoding GTP-binding protein from a fish cell, CHSE-214. The clone was 1493 bp long and contained an open reading frame encoding 364 amino acids. It has the five sequence motifs G1-G5 that are conserved in all GTP-binding proteins. Its amino acid sequences are strikingly different from those of the well-characterized G-proteins. However, sequences closely related to this protein are found in various kinds of species including human, Arabidopsis, Drosophila and archaebacteria, suggesting a novel subfamily within the superfamily of the GTP-binding proteins. Northern analysis indicates that this gene is constitutively expressed at a low level in normal cells but is induced by fish rhabdovirus infection at about 24 h post infection and disappears thereafter. Based on these observations, we propose that this protein represents an evolutionarily conserved novel subfamily of GTP-binding proteins which may play an important role in fish rhabdovirus infection.

Analysis of mutations of neurofibromatosis type 1 gene and N-ras gene in acute myelogenous leukemia.

Neurofibromatosis type 1 (NF1) gene is a tumor suppressor gene, and the NF1 gene product, neurofibromin, can downregulate the N-ras gene. Because the N-ras gene is often mutated in acute myelogenous leukemia (AML), we wondered if the NF1 gene might be mutated in those AML samples not having N-ras mutations. We investigated the mutational status of the N-ras gene and the FLR exon of codons 1371-1423 of the open reading frame of the full-length NF1 cDNA, which has a strong homology with the mammalian ras GTPase-activating protein (GAP), especially for a stretch of three consecutive amino acids (F, L, R), by single-strand conformation polymorphism analysis and direct sequencing in samples from patients with AML. Of 48 AML patients, 10 (21%) had point (missense) mutations of the N-ras gene involving codons 12, 13 and 61. However, mutations in the FLR exon of the NF1 gene were not detected in any of the AML samples. We also examined the difference of clinical response to induction therapy between AML patients with and without N-ras mutation. A significantly lower rate of complete remission was noted in individuals with N-ras gene mutations. These results suggest that mutation of the NF1 gene, at least in the FLR exon, is very rare in AML and the NF1 gene probably is not a functional complement of the N-ras gene mutation. The presence of N-ras gene mutation may be associated with a lower clinical response to antileukemic therapy.

Testicular neoplasms in Hispanic brothers.

Two hispanic brothers developed nonconcordant testicular malignancies in an interval of 11 months. They represent the 36th report of familial testicular neoplasia. The problems deriving from poor patient compliance are documented.

Malignant feminizing Leydig cell tumor.

A patient with a feminizing malignant Leydig cell tumor is presented. Hormonal assays revealed increased production of prolactin, estradiol, and total estrogens. Eleven years after the onset of his disease he remains clinically well.