Compressive stress induces collective migration through cytoskeletal remodelling in nasal polyp epithelium.

BACKGROUND: Nasal polyps in the nasal cavity and mucous discharge inside the maxillary sinus exhibit compressive stress on the nasal mucosal epithelium. However, there have been only a few studies on how compressive stress impacts the human nasal mucosal epithelium. METHODOLOGY: We investigated the effect of compressive stress on collective migration, junctional proteins, transepithelial electri- cal resistance, epithelial permeability, and gene expression in well-differentiated normal human nasal epithelial (NHNE) cells and human nasal polyp epithelial (HNPE) cells. RESULTS: NHNE cells barely showed collective migration at compressive stress up to 150 mmH20. However, HNPE cells showed much greater degree of collective migration at a lower compressive stress of 100 mmH20. The cell migration of HNPE cells sub- jected to 100 mmH2O compression was significantly decreased at day 3 and was recovered to the status prior to the compressive stress by day 7, indicating that HNPE cells are relatively more sensitive to mechanical pressure than NHNE cells. Compressive stress also increased transepithelial electrical resistance and decreased epithelial permeability, indicating that the compressive stress disturbed the structural organization rather than physical interactions between cells. In addition, we found that compressive stress induced gene expressions relevant to airway inflammation and tissue remodelling in HNPE cells. CONCLUSION: Taken together, these findings demonstrate that compressive stress on nasal polyp epithelium is capable of inducing collective migration and induce increased expression of genes related to airway inflammation, innate immunity, and polyp remo- delling, even in the absence of inflammatory mediators.

Red Blood Cell Distribution Width Is Associated with Carotid Atherosclerosis in People with Type 2 Diabetes.

AIMS: Red cell distribution width (RDW) has been shown to be associated with cardiovascular diseases (CVD). The relationship between RDW and carotid intima-media thickness (C-IMT), a marker of subclinical atherosclerosis, has been inconsistent in subjects with cardiovascular risk factors. In this study, we investigated the relationship between RDW and carotid atherosclerosis in people with type 2 diabetes. METHODS: Four hundred sixty-nine people with type 2 diabetes without history of cardiovascular or cerebrovascular diseases were enrolled. Anthropometric measures and various biochemical parameters including RDW were assessed. Ultrasonographic measurement of carotid intima-media thickness was used to evaluate subclinical atherosclerosis. RESULTS: The participants were stratified into 3 groups according to RDW. The C-IMT increased gradually according to RDW tertiles (lowest, second, highest RDW tertiles; 0.740 ± 0.120, 0.772 ± 0.138, and 0.795 ± 0.139, respectively; p < 0.01). Multiple regression analysis and multivariate logistic regression analysis revealed that RDW was associated with C-IMT in people with type 2 diabetes, and it remained significant after control for various cardiovascular risk factors including body mass index, blood pressure, insulin resistance, and smoking status in multivariate logistic regression analysis. CONCLUSION: RDW is associated with subclinical atherosclerosis assessed by carotid IMT after control of various covariates in people with type 2 diabetes without cardiovascular or cerebrovascular diseases.

Apoptotic effect of dibenzylideneacetone on oral cancer cells via modulation of specificity protein 1 and Bax.

OBJECTIVE: Dibenzylideneacetone (DBA), an analogue of curcumin, has been shown to have potential anticancer effects against several cancers. However, the molecular mechanism underlying anticancer activity of DBA has not been well established yet. In this study, we investigated the function and molecular mechanism of DBA in human oral cancer cells. MATERIALS AND METHODS: The growth-inhibitory and apoptotic effects and related signaling pathways of DBA were evaluated using trypan blue exclusion assay, 4'-6-diamidino-2-phenylindole staining, Western blot analysis, siRNA, and reverse transcription-polymerase chain reaction. RESULTS: DBA inhibited cell growth and induced apoptosis, as evidenced by PARP cleavage, activation of caspase-3, and nuclear condensation. DBA also decreased specificity protein 1 (Sp1) expression through facilitating protein degradation. In addition, DBA enhanced the induction of pro-apoptotic protein Bax, resulting in their conformational change, translocation into mitochondrial outer membrane, and its oligomerization. The down-regulation of Sp1 by siRNA targeting Sp1 and mithramycin A increasingly activated Bax to trigger apoptosis. Moreover, DBA-induced growth inhibition and apoptosis in various human oral cancer cell lines were associated with Sp1 down-regulation and induction of Bax. CONCLUSION: These findings suggest that DBA may be a potential anticancer drug candidate to induce apoptosis through down-regulation of Sp1 in human oral cancer.

The humoral immune response to the inactivated influenza A (H1N1) 2009 monovalent vaccine in patients with Type 2 diabetes mellitus in Korea.

AIMS: We evaluated the antibody response to a single-dose adjuvanted, inactivated, pandemic H1N1 influenza vaccination in patients with diabetes and assessed factors associated with the failure to induce antibody responses. METHODS: Eighty-two patients with Type 2 diabetes were vaccinated and antibody responses were determined with haemagglutination inhibition assay and anti-haemagglutinin antibody ELISA. RESULTS: Among 70 antibody-negative patients at baseline, 34 (48.6%) achieved seroconversion; 28 (60.9%) in the young adults group and six (25%) in the elderly group acquired H1N1-specific antibodies. Patients in the older age range or with longer duration of diabetes had a lower seroconversion rate. CONCLUSIONS: Our data show low cross-reactive antibody carrying rate and low seroconversion rate in patients with diabetes. Until larger-scale, case-controlled trials become available, older patients and patients with a longer duration of diabetes should be considered for the two-dose vaccination or have antibody titres measured after the first vaccination.

Tribological property of the cobalt-chromium femoral head with different regions of wear in total hip arthroplasty.

The tribological properties of engineering and biological materials have been investigated at microscale levels through the calculation of the surface roughness and frictional coefficient using atomic force microscopy (AFM). Although a number of previous studies have reported the frictional coefficients of diverse bearing materials in total hip arthroplasty (THA), the relationship between the surface roughness and frictional coefficient of bearing materials of THA have not been reported, and furthermore, the tribological properties for different wear regions of a cobalt-chromium (Co-Cr) femoral head have not been well identified. Therefore, the objective of this study is to investigate the relationships between the surface roughness, frictional coefficient, and hardness for both the main-wear and the least-wear regions of a Co-Cr femoral head 10 years after THA. The average Vickers hardness of the Co-Cr femoral head was 380.7 +/- 11.3 HV. With the scanned area of 25 microm x 25 microm through AFM, the frictional coefficients of the main-wear and the least-wear regions were 0.229 +/- 0.054 and 0.243 +/- 0.059, respectively, and showed no statistical differences between these two regions (p = 0.449). However, differences in the surface roughness (Rq) between the main-wear region (Rq = 96.5 +/- 26.2 nm) and the least-wear region (Rq = 17.7 +/- 4.2 nm) were statistically significant (p < 0.0001). The results of the current study suggest that the frictional property of the Co-Cr femoral head is not significantly correlated with its surface roughness, and also provide guidelines for improving the surface characteristics of metallic implant materials.

The effect of rosiglitazone on insulin sensitivity and mid-thigh low-density muscle in patients with Type 2 diabetes.

AIMS: We examined the effect of rosiglitazone on insulin sensitivity, abdominal fat and mid-thigh intramuscular fat distribution, and plasma concentrations of adipocytokines in patients with Type 2 diabetes. METHODS: Rosiglitazone was administered at a daily dose of 4 mg to 42 Type 2 diabetes patients [age 32-70 years, body mass index (BMI) 17.5-32.6 kg/m(2), 15 women, 27 men] for 12 weeks. Various anthropometric and metabolic profiles, plasma adiponectin, leptin, and resistin levels were measured, and insulin resistance was calculated from the short insulin tolerance test. Body fat composition was assessed by computed tomography. RESULTS: Twelve weeks' rosiglitazone treatment resulted in improved insulin resistance despite increases in body weight and BMI. There was a significant decrease in abdominal visceral adipose tissue area (145 +/- 65.6 vs. 129 +/- 73.1 cm(2), P = 0.049). Mid-thigh low-density muscle area (TLDMA) increased from 23 +/- 9.6 to 26 +/- 8.2 cm(2) (P = 0.009). There were significant changes in plasma adipocytokines, but they were not significantly correlated with changes in insulin resistance. CONCLUSIONS: Rosiglitazone treatment resulted in an improvement of insulin responsiveness in Type 2 diabetic subjects, which was associated with the redistribution of visceral and subcutaneous adipose tissue, an increase in TLDMA, and changes in serum adipocytokine levels. Further studies are needed to elucidate the insulin sensitizing mechanism of rosiglitazone on peripheral skeletal muscles.

The effect of mosapride (5HT-4 receptor agonist) on insulin sensitivity and GLUT4 translocation.

AIMS: We investigated the effect of mosapride, 5HT-4 (5-hydroxytryptamine) agonist, on blood glucose level and insulin sensitivity in subjects with impaired glucose tolerance (IGT) and conducted an in vitro study to evaluate the action mechanism. METHODS: Thirty IGT patients were randomly assigned to receive either mosapride or placebo for 2 weeks. Biochemical profiles and insulin sensitivity index from euglycemic hyperinsulinemic clamp test were assessed before and after treatment. In cultured myotubes from human skeletal muscle cells, insulin- and mosapride-induced GLUT4 translocation and tyrosine phosphorylation of IRS-1 were determined. RESULTS: After 2 weeks of treatment with mosapride, glucose disposal rates were significantly increased up to those of control (mosapride 5.47+/-1.72 vs 7.06+/-2.13, P=0.004, placebo 5.42+/-1.85 vs 5.23+/-1.53mgkg(-1)min(-1)). Fasting plasma glucose (FPG) and insulin levels were decreased. Mosapride increased the contents of GLUT4 in plasma membrane representing the increased recruitment of glucose transporters from intracellular pool. While insulin treatment on human skeletal muscle cell resulted in an increased tyrosine phosphorylation of IRS-1, mosapride did not have any effect. CONCLUSIONS: Mosapride is effective in decreasing FPG without stimulating insulin secretion in IGT subjects, possibly by inducing GLUT4 translocation in skeletal muscles.

Nerve growth factor and expression of its receptors in patients with diabetic neuropathy.

AIMS: Low serum nerve growth factor (NGF) levels have been reported in patients with diabetic peripheral neuropathy (DPN), but the role of NGF in the development of neuropathy is unclear. Thus, we investigated the associations of serum NGF level and NGF receptor activity with the presence and severity of DPN. METHODS: One hundred and thirty-six patients with Type 2 diabetes were included in this cross-sectional study. Serum NGF levels were measured by ELISA. Expressions of NGF receptors (TrkA and p75(NTR)) were measured by immunohistochemical staining. The presence and severity of DPN were assessed by neuropathy disability score (NDS) and by corneal nerve fibre length (cNFL) and nerve branch density (cNBD) using in vivo confocal microscopy. RESULTS: Patients with DPN had higher serum NGF levels (56-451 pg/ml) than patients without DPN (4-54 pg/ml). However, in DPN patients, serum NGF was negatively associated with neuropathy severity (mild 222 +/- 64 pg/ml; moderate 114 +/- 17 pg/ml; severe 89 +/- 20 pg/ml). This negative association was consistent in all severity indices (NDS, P < 0.001; cNFL, P < 0.001; cNBD P = 0.010) even after adjustment for age, sex, diabetes duration, insulin use, fasting glucose and glycated haemoglobin. Although NGF receptor activities had significantly (P < 0.05) negative associations with the presence and severity of neuropathy, these associations were not significant when adjusted for other factors. CONCLUSIONS: Serum NGF level was positively associated with the presence of DPN but negatively associated with neuropathy severity in DPN patients. The change in serum NGF might be a consequence of, rather than a contributor to, the early development of DPN.

The association between pulse wave velocity and metabolic syndrome and adiponectin in patients with impaired fasting glucose: cardiovascular risks and adiponectin in IFG.

We aimed to assess how metabolic profiles, surrogate markers of insulin resistance, and subclinical atherosclerosis are interrelated in subjects with impaired fasting glucose (IFG) and investigate whether the diagnosis of metabolic syndrome (MetS) further increases the risk of cardiovascular disease among subjects already at risk. We analyzed 1739 Korean subjects with IFG. The parameters of MetS, plasma adiponectin level, and pulse wave velocity (PWV) were assessed. Subjects with MetS had unfavorable metabolic parameters, lower adiponectin level, and higher peripheral PWV compared to those without MetS. Adiponectin correlated with fasting glucose, waist circumference, triglyceride, HDL-cholesterol, BMI, HOMA-IR, and the number of MetS components. In addition to blood pressure, peripheral PWV was associated with triglyceride, waist circumference, and the number of MetS components while aortic PWV correlated positively with fasting plasma glucose. Multiple linear regression analysis revealed that adiponectin correlated with HDL-cholesterol, HOMA-IR, fasting glucose, waist circumference, and triglyceride, peripheral PWV with blood pressure, body mass index, waist circumference, and the number of MetS components, and aortic PWV with fasting plasma glucose. In subjects with IFG, concurrent MetS increases PWV and has an unfavorable effect on cardiovascular risks, and these risks were further increased by additional MetS components.

Cryopreserved human adipogenic-differentiated pre-adipocytes: a potential new source for adipose tissue regeneration.

BACKGROUND: Previously, we have shown that in vitro adipogenic differentiation of pre-adipocytes before implantation can enhance in vivo adipose tissue formation. For large-scale adipose tissue engineering or repeat procedures, cryopreservation of fat grafts has been commonly used in recent years. However, the feasibility of cryopreservation of adipogenic differentiated pre-adipocytes has not been investigated. METHODS: To examine the impact of cryopreservation on the adipogenic functions of adipogenic-differentiated pre-adipocytes, freeze-thawed adipocytes were compared with fresh differentiated adipocytes in vitro and in vivo. Adipogenic function was assessed by Oil red O staining, ELISA analysis of leptin secretion and RT-PCR of adipogenic-related genes. After transplantation, adipose tissue formation was assessed by histomorphologic and volumetric analysis. RESULTS: Freeze-thawed adipocytes constantly showed typical adipogenic functions in terms of lipid content, leptin secretion and adipogenic gene expression, as well as good viability. Importantly, implants derived from freeze-thawed adipocytes were successfully developed to adipose tissue and newly formed adipose tissues were similar to those developed from fresh differentiated adipocytes, based on histomorphologic and volumetric analysis. In addition, CD34-positive endothelial cells were detected in implants. These results demonstrate that the specific characters of adipogenic-differentiated pre-adipocytes are successfully conserved after cryopreservation without any significant alteration. DISCUSSION: Cryopreservation of adipogenic-differentiated pre-adipocytes is a feasible method and extends their clinical use in adipose tissue-engineering applications and transplantation.

The clinical and immunogenetic characteristics of adult-onset type 1 diabetes mellitus in Korea.

Although the HLA class II alleles and immunological abnormalities are associated with type 1 diabetes mellitus (T1DM) in all racial groups, there are considerable variations in the genotypes and the prevalence of autoantibodies. In order to investigate the characteristics of the immunogenetic patterns and to use these as an early diagnostic tool and guideline for a therapeutic plan, we examined the clinical characteristics and the patterns of anti-GAD antibody (GADA), IA-2 antibody (IA-2A), HLA-DR and HLA-DQ in Korean adult-onset T1DM patients. Adult-onset patients had higher serum C-peptide levels than child-onset patients. In adult-onset patients, the prevalence of GADA and IA-2A were 59.5% and 15.3% respectively, and increased frequencies of HLADR4 and-DR9 were found. The frequencies of HLADQA1,-DQB1 and-DQ heterodimers were similar to those of the control, but child-onset patients had high frequencies of the HLA-DR3,-DR4,-DR9, DQA1*0301, DQA1*0501 and DQB1*0201 genotypes. In conclusion, Korean adult-onset T1DM patients had a lower prevalence of GADA, which was comparable to that found in Caucasian patients. The detection of GADA might help to predict the insulin dependency of adult-onset diabetes. Difference in the frequencies of diabetes associated with HLA type suggests that there might be a heterogeneity in the pathogenesis of diabetes according to the age of onset.

AtBAG6, a novel calmodulin-binding protein, induces programmed cell death in yeast and plants.

Calmodulin (CaM) influences many cellular processes by interacting with various proteins. Here, we isolated AtBAG6, an Arabidopsis CaM-binding protein that contains a central BCL-2-associated athanogene (BAG) domain. In yeast and plants, overexpression of AtBAG6 induced cell death phenotypes consistent with programmed cell death (PCD). Recombinant AtBAG6 had higher affinity for CaM in the absence of free Ca2 + than in its presence. An IQ motif (IQXXXRGXXXR, where X denotes any amino-acid) was required for Ca2 +-independent CaM complex formation and single amino-acid changes within this motif abrogated both AtBAG6-activated CaM-binding and cell death in yeast and plants. A 134-amino-acid stretch, encompassing both the IQ motif and BAG domain, was sufficient to induce cell death. Agents generating oxygen radicals, which are known to be involved in plant PCD, specifically induced the AtBAG6 transcript. Collectively, these results suggest that AtBAG6 is a stress-upregulated CaM-binding protein involved in plant PCD.

Srg3, a mouse homolog of yeast SWI3, is essential for early embryogenesis and involved in brain development.

Srg3 (SWI3-related gene product) is a mouse homolog of yeast SWI3, Drosophila melanogaster MOIRA (also named MOR/BAP155), and human BAF155 and is known as a core subunit of SWI/SNF complex. This complex is involved in the chromatin remodeling required for the regulation of transcriptional processes associated with development, cellular differentiation, and proliferation. We generated mice with a null mutation in the Srg3 locus to examine its function in vivo. Homozygous mutants develop in the early implantation stage but undergo rapid degeneration thereafter. An in vitro outgrowth study revealed that mutant blastocysts hatch, adhere, and form a layer of trophoblast giant cells, but the inner cell mass degenerates after prolonged culture. Interestingly, about 20% of heterozygous mutant embryos display defects in brain development with abnormal organization of the brain, a condition known as exencephaly. Histological examination suggests that exencephaly is caused by the failure in neural fold elevation, resulting in severe brain malformation. Our findings demonstrate that Srg3 is essential for early embryogenesis and plays an important role in the brain development of mice.

Kitasatospora cheerisanensis sp. nov., a new species of the genus Kitasatospora that produces an antifungal agent.

An actinomycete, strain YC75T, which produced bafilomycin-like antifungal compounds, was identified as a member of the genus Kitasatospora on the basis of morphological and chemotaxonomic characteristics. The strain produced the aerial and fragmenting vegetative mycelia consisting of straight chains of 20 or more smooth-surfaced spores. Submerged spores were formed in tryptic soy broth. No soluble pigments were formed. Whole-cell hydrolysates contained glucose and mannose, but not galactose. The 16S rDNA sequence of YC75T was compared with those of the other representative kitasatosporae and streptomycetes. Strain YC75T formed a significant monophyletic clade with Kitasatospora phosalacinea. The levels of DNA relatedness between strain YC75T and representatives of the genus Kitasatospora ranged from 16 to 59% including K. phosalacinea (28 and 40%). It is clear from polyphasic evidence that the isolate should be classified as Kitasatospora cheerisanensis sp. nov., whose type strain is YC75T (= KCTC 2395T). The presence of galactose in whole-cell hydrolysates may not be a stable chemical marker for the genus Kitasatospora.

Cloning and phylogenetic analysis of chitin synthase genes from the insect pathogenic fungus, Metarhizium anisopliae var. anisopliae.

Degenerated PCR primers were used to amplify chitin synthase genes from genomic DNA of Metarhizium anisopliae var. anisopliae. Through cloning and sequencing of approximately 600-bp fragments amplified by PCR, we found three genes encoding different types of chitin synthases, designated MaCHS1, MaCHS2, and MaCHS3. Southern blot analysis performed on genomic DNA showed that each of the chitin synthases MaCHS1, MaCHS2, and MaCHS3 is encoded by a single copy gene. Alignment of their deduced amino acid sequences with those of other euascomycetes separated the sequences into three distinct classes. MaCHS1 was identified as a gene for class I chitin synthase, MaCHS2 for class II, and MaCHS3 for class III. The UPGMA dendrogram and phylogenetic tree of the deduced amino acid sequences revealed the taxonomic and evolutionary position of Metarhizium anisopliae var. anisopliae.

A protein kinase C-activating phorbol ester enhances transcription of the human DBH gene through a cyclic AMP response element in SK-N-BE(2)C cells.

Protein kinase C (PKC) activation after treatment of human neuroblastoma SK-N-BE(2)C cells with phorbol 12-myristate 13-acetate (PMA) was found to enhance transcription of the human dopamine beta-hydroxylase (DBH) in those cells. To identify which cis-acting element is responsive to the PMA treatment during DBH gene expression, we employed transient transfection assays with serially deleted constructs of the human DBH gene's 5' upstream region fused to the chloramphenicol acetyltransferase (CAT) gene. Treatment of transfected cells with PMA resulted in an approximate threefold increase in CAT expression for all deletion constructs ranging from -978 bp to -262 bp, while the enhancement did not occur with a construct shortened to -114 bp. The region between -262 and -114 bp from the initiation site of transcription contains several cis-regulatory elements including a cyclic AMP response element (CRE) and putative AP1 and YY1 sequences. Site-directed mutagenesis of those cis-acting elements were performed to identify which of the elements mediated the PMA-induced transcriptional enhancement. Substitution of bases in the putative AP1 site containing in part a putative YY1 sequence did not effect the PMA inducibility. However, specific mutations in the CRE sequence abolished the PMA-inducible effect. Changing the CRE sequence into an authentic AP1 sequence (TGACGTCC --> TGACTCA) did not affect the PMA inducibility, suggesting that AP1 factors might interact with the new AP1 site upon PKC activation. A specific PKC inhibitor, GF109203X, completely inhibited the stimulatory effect of PMA on the expression of the human DBH gene. PMA induced an increase in the DBH mRNA level as detected by Northern blot analysis. Gel retardation showed that the binding of nuclear factors to CRE, putative YY1, and AP1 was sequence specific. Our data suggest that the enhancement of the human DBH gene expression by PMA treatment is mediated by the CRE motif in the 5' upstream region of the gene, and occurs via a PKC-dependent pathway.

Bioflocculant produced by Aspergillus sp. JS-42.

A bioflocculant from a fungus, Aspergillus sp. JS-42, was purified by precipitations with acetone and cetylpyridinium chloride. The flocculating activity was not significantly affected by pH from 3 to 8, but was stimulated by the addition of CaCl2, and was effective only when the reaction mixture contained an adequate amount of flocculant. The flocculant could efficiently flocculate all tested solids suspended in aqueous solution, including various microorganisms, organic acids, and inorganic materials.