Caveolin-1 abolishment attenuates the myogenic response in murine cerebral arteries

Intravascular pressure-induced vasoconstriction (the "myogenic response") is intrinsic to smooth muscle cells, but mechanisms that underlie this response are unresolved. Here we investigated the physiological function of arterial smooth muscle cell caveolae in mediating the myogenic response. Since caveolin-1 (cav-1) ablation abolishes caveolae formation in arterial smooth muscle cells, myogenic mechanisms were compared in cerebral arteries from control (cav-1(+/+)) and cav-1-deficient (cav-1(-/-)) mice. At low intravascular pressure (10 mmHg), wall membrane potential, intracellular calcium concentration ([Ca(2+)](i)), and myogenic tone were similar in cav-1(+/+) and cav-1(-/-) arteries. In contrast, pressure elevations to between 30 and 70 mmHg induced a smaller depolarization, [Ca(2+)](i) elevation, and myogenic response in cav-1(-/-) arteries. Depolarization induced by 60 mM K(+) also produced an attenuated [Ca(2+)](i) elevation and constriction in cav-1(-/-) arteries, whereas extracellular Ca(2+) removal and diltiazem, an L-type Ca(2+) channel blocker, similarly dilated cav-1(+/+) and cav-1(-/-) arteries. N(omega)-nitro-l-arginine, an nitric oxide synthase inhibitor, did not restore myogenic tone in cav-1(-/-) arteries. Iberiotoxin, a selective Ca(2+)-activated K(+) (K(Ca)) channel blocker, induced a similar depolarization and constriction in pressurized cav-1(+/+) and cav-1(-/-) arteries. Since pressurized cav-1(-/-) arteries are more hyperpolarized and this effect would reduce K(Ca) current, these data suggest that cav-1 ablation leads to functional K(Ca) channel activation, an effect that should contribute to the attenuated myogenic constriction. In summary, data indicate that cav-1 ablation reduces pressure-induced depolarization and depolarization-induced Ca(2+) influx, and these effects combine to produce a diminished arterial wall [Ca(2+)](i) elevation and constriction.     

RAPD typing of Aspergillus chevalieri, Aspergillus nidulans, Aspergillus tetrazonus (quadrilineatus) and their teleomorphs using 5'-d[AACGCGCAAC]-3' and 5'-d[CCCGTCAGCA]-3' primers

Sexuality in fungi has long been a matter of concerns and debates that always necessitated extensive analysis of the relationship between organisms assumed to represent different developmental forms of the same organism. Moreover, mating-virulence correlation and the growing worry of opportunistic fungal infections in immunocompromised states associated with AIDS, cancer chemotherapy and organ transplantation protocols have been critically addressed nowadays. In view of that, we genetically characterized Aspergillus chevalieri, Aspergillus nidulans, Aspergillus tetrazonus (quadrilineatus) and their teleomorphs with RAPD analysis using 5'-d[AACGCGCAAC]-3' and 5'-d[CCCGTCAGCA]-3' primers. The reported similarities between the sexual and the asexual forms of the tested species, using Dice coefficient, ranged between 40% and 70% which holds pretty much with the current systematics of the genus. The study presents a rapid consistent method for identification of A. chevalieri, A. nidulans, A. tetrazonus (quadrilineatus) and their teleomorphs based on the banding pattern of RAPD-generated fragments that can be reliably used ahead of further applications on these species.     

Modelling the population dynamics of <Emphasis Type="Italic">Daphnia obtusa</Emphasis> (Kurz) in Lake Orta (N. Italy) under pre- and post-liming conditions

A mathematical matrix model was formulated to investigate the response of Daphnia obtusa population dynamics to the changes in the water chemistry of Lake Orta before and after the liming operation. Model parameters were estimated from experimental laboratory data. Model analysis showed that water chemistry changes induced by liming affected mainly egg survival and predicted the highest population growth at pH␣6. Whereas increased egg mortality heavily inhibits population growth rate, the model still predicts a long term tendency of the population to increase in number. However, both before and after the liming operation due to high food availability in the laboratory, egg production was higher under all experimental conditions than in the field. When food limitation is accounted for and more realistic, field based estimates of egg production are used, the model predicts the extinction of D. obtusa population in the lake. This suggests that the effects of water chemistry changes on egg mortality had a critical role in the disappearance of D. obtusa from Lake Orta and may even adequately explain the extinction of this population.     

Investigation of Thermal and Viscoelastic Properties of Polymers Relevant to Hot Melt Extrusion,IV: Affinisol™ HPMC HME Polymers

Most cellulosic polymers cannot be used as carriers for preparing solid dispersion of drugs by hot melt extrusion (HME) due to their high melt viscosity and thermal degradation at high processing temperatures. Three HME-grade hydroxypropyl methylcelluloses, namely Affinisol™ HPMC HME 15 cP, Affinisol™ HPMC HME 100 cP, and Affinisol™ HPMC HME 4 M, have recently been introduced by The Dow Chemical Co. to enable the preparation of solid dispersion at lower and more acceptable processing temperatures. In the present investigation, physicochemical properties of the new polymers relevant to HME were determined and compared with that of Kollidon^® VA 64. Powder X-ray diffraction (PXRD), modulated differential scanning calorimetry (mDSC), thermogravimetric analysis (TGA), moisture sorption, rheology, and torque analysis by melt extrusion were applied. PXRD and mDSC showed that the Affinisol™ polymers were amorphous in nature. According to TGA, the onset of degradation for all polymers was >220°C. The Affinisol™ polymers exhibited less hygroscopicity than Kollidon^® VA 64 and another HPMC polymer, Methocel™ K100LV. The complex viscosity profiles of the Affinisol™ polymers as a function of temperature were similar. The viscosity of the Affinisol™ polymers was highly sensitive to the shear rate applied, and unlike Kollidon^® VA 64, the viscosity decreased drastically when the angular frequency was increased. Because of the very high shear rate encountered during melt extrusion, Affinisol™ polymers showed capability of being extruded at larger windows of processing temperatures as compared to that of Kollidon^® VA 64.KEY WORDS: Affinisol™ HPMC HME, hot melt extrusion, hydroxypropyl methylcellulose, solid dispersion, thermal analysis, viscosity     

MRSA Clonal Complex 22 Strains Harboring Toxic Shock Syndrome Toxin (TSST-1) Are Endemic in the Primary Hospital in Gaza,Palestine

Background

Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in both community and healthcare-related settings worldwide. Current knowledge regarding the epidemiology of S. aureus and MRSA in Gaza is based on a single community-based carriage study. Here we describe a cross-sectional analysis of 215 clinical isolates collected from Al-Shifa Hospital in Gaza during 2008 and 2012.

Methods

All isolates were characterized by spa typing, SCCmec typing, and detection of genes encoding Panton-Valentine leukocidin (PVL) and toxic shock syndrome toxin (TSST-1). Representative genotypes were also subjected to multilocus sequence typing (MLST). Antibiotic susceptibility testing was performed using VITEK2 and MicroScan.

Results

MRSA represented 56.3% of all S. aureus strains, and increased in frequency from 2008 (54.8%) to 2012 (58.4%). Aside from beta-lactams, resistance was observed to tetracycline, erythromycin, clindamycin, gentamicin, and fluoroquinolones. Molecular typing identified 35 spa types representing 17 MLST clonal complexes (CC), with spa 998 (Ridom t223, CC22) and spa 70 (Ridom t044, CC80) being the most prevalent. SCCmec types I, III, IV, V and VI were identified among MRSA isolates, while type II was not detected. PVL genes (lukF/S-PV) were detected in 40.0% of all isolates, while the TSST-1 gene (tst) was detected in 27.4% of all isolates, with surprisingly high frequency within CC22 (70.4%). Both PVL and TSST-1 genes were found in several isolates from 2012.

Conclusions

Molecular typing of clinical isolates from Gaza hospitals revealed unusually high prevalence of TSST-1 genes among CC22 MRSA, which is noteworthy given a recent community study describing widespread carriage of a CC22 MRSA clone known as the ‘Gaza strain’. While the latter did not address TSST-1, tst-positive spa 998 (Ridom t223) has been detected in several neighboring countries, and described as endemic in an Italian NICU, suggesting international spread of a ‘Middle Eastern variant’ of pandemic CC22 strain EMRSA-15.     

MicroRNA Profiling in Intraocular Medulloepitheliomas

Purpose

To study the differential expression of microRNA (miRNA) profiles between intraocular medulloepithelioma (ME) and normal control tissue (CT).

Material and Methods

Total RNA was extracted from formalin fixed paraffin embedded (FFPE) intraocular ME (n=7) and from age matched ciliary body controls (n=8). The clinical history and phenotype was recorded. MiRNA profiles were determined using the Affymetrix GeneChip miRNA Arrays analyzed using expression console 1.3 software. Validation of significantly dysregulated miRNA was confimed by quantitaive real-time PCR. The web-based DNA Intelligent Analysis (DIANA)-miRPath v2.0 was used to perform enrichment analysis of differentially expressed (DE) miRNA gene targets in Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway.

Results

The pathologic evaluation revealed one benign (benign non-teratoid, n=1) and six malignant tumors (malignant teratoid, n=2; malignant non-teratoid, n = 4). A total of 88 miRNAs were upregulated and 43 miRNAs were downregulated significantly (P<0.05) in the tumor specimens. Many of these significantly dysregulated miRNAs were known to play various roles in carcinogenesis and tumor behavior. RT-PCR validated three significantly upregulated miRNAs and three significantly downregulated miRNAs namely miR-217, miR-216a, miR-216b, miR-146a, miR-509-3p and miR-211. Many DE miRNAs that were significant in ME tumors showed dysregulation in retinoblastoma, glioblastoma, and precursor, normal and reactive human cartilage. Enriched pathway analysis suggested a significant association of upregulated miRNAs with 15 pathways involved in prion disease and several types of cancer. The pathways involving significantly downregulated miRNAs included the toll-like receptor (TLR) (p<4.36E-16) and Nuclear Factor kappa B (NF-κB) signaling pathways (p<9.00E-06).

Conclusions

We report significantly dysregulated miRNAs in intraocular ME tumors, which exhibited abnormal profiles in other cancers as well such as retinoblastoma and glioblastoma. Pathway analysis of all dysregulated miRNAs shared commonalities with other cancer pathways.     

Protective effect of proanthocyanidins against doxorubicin‐induced nephrotoxicity in rats

Doxorubicin (DOX) exerts toxic effects in several organs particularly kidney. The present study aimed to assess the protective effect of proanthocyanidins (PAs) against DOX‐induced nephrotoxicity in rats. A single dose of DOX (7.5 mg/kg, i.v.) significantly increased kidney weight, kidney/body weight ratio, serum urea, creatinine, tumor necrosis factor alpha levels, and kidney contents of malondialdehyde, nitric oxide, cyclooxygenase‐2, and caspase‐3 activity with significant reduction in final body weight, serum albumin, kidney contents of reduced glutathione (GSH), and superoxide dismutase activity as compared with control group. In contrast, pretreatment with PAs (200 mg/kg, p.o.) for 14 days before DOX and for 7 days after DOX ameliorated kidney function and oxidative stress parameters. Histopathological evidence confirmed the protective effects of PAs from the tissue damage induced by DOX. In conclusion, PAs have a multi‐nephroprotective effect that might be attributed to its antioxidant, anti‐inflammatory, and antiapoptoic activities.     

Evidence for Zoonotic Potential of Enterocytozoon bieneusi in Its First Molecular Characterization in Captive Mammals at Bangladesh National Zoo

To determine the occurrence and genotypes of Enterocytozoon bieneusi in captive mammals at Bangladesh National Zoo and to assess their zoonotic significance, 200 fecal samples from 32 mammalian species were examined using a nested PCR and sequencing of internal transcribed spacer (ITS) gene. Enterocytozoon bieneusi was detected in 16.5% (33/200) of the samples. Seven different ITS genotypes were identified, including two known genotypes (D and J) and five new ones (BAN4 to BAN8). Genotype D was the most common genotype being observed in 19 isolates. In phylogenetic analysis, four genotypes (D, BAN4, BAN5, and BAN6), detected in 30 isolates (90.9%), belonged to Group 1 having zoonotic potential. The sequence of genotype J found in a Malayan pangolin was clustered in so‐called ruminant‐specific Group 2. The other two genotypes BAN7 and BAN8 were clustered in primate‐specific Group 5. To our knowledge, this is the first report of molecular characterization of E. bieneusi in Bangladesh, particularly in captive‐bred wildlife in this country. The potentially zoonotic genotypes of E. bieneusi are maintained in zoo mammals that may transmit among these animals and to the humans through environmental contamination or contact.     

Clinical Characteristics,Etiology and Antimicrobial Susceptibility among Overweight and Obese Individuals with Diarrhea: Observed at a Large Diarrheal Disease Hospital,Bangladesh

Background

The present study aimed to determine the clinical characteristics and etiology of overweight and obese (OO) individuals with diarrhea attending an urban Dhaka Hospital, International Centre for Diarrheal Disease Research (icddr,b), Bangladesh.

Methods

Total of 508 under-5 children, 96 individuals of 5–19 years and 1331 of >19 years were identified as OO from the Diarrheal Disease Surveillance System (DDSS) between 1993–2011. Two comparison groups such as well-nourished and malnourished individuals from respective age stratums were selected.

Results

Isolation rate of rotavirus was higher among OO under-5 children compared to malnourished group (46% vs. 28%). Rotavirus infection among OO individuals aged 5–19 years (9% vs. 3%) (9% vs. 3%) and >19 years (6% vs. 4%) (6% vs. 3%) was higher compared to well-nourished and malnourished children. Conversely, Vibrio cholerae was lower among all OO age groups compared to well-nourished and malnourished ones. Shigella (4% vs. 6%) (4% vs. 8%), and Campylobacter (3% vs. 5%) (3% vs. 5%) were lower only among OO in >19 years individuals compared to their counterparts of the same age stratum. Salmonella was similarly isolated in all age strata and nutritional groups. In multinomial logistic regression among under-5 children, significant association was observed only with use of antimicrobials at home [OR-1.97] and duration of hospital stay [OR-0.68]. For individuals aged 5–19 years, use of antimicrobials at home (OR-1.83), some or severe dehydration (OR-3.12), having received intravenous saline (OR-0.46) and rotavirus diarrhea (OR-2.96) were found to be associated with OO respectively. Moreover, significant associations were also found for duration of diarrhea before coming to hospital (>24 hours) (OR-1.24), Shigella (OR-0.46), and Campylobacter (OR-0.58) among >19 years OO individuals along with other associated co-variates in 5–19 years group (all p<0.05).

Conclusion and significance

Higher proportion of OO were infected with rotavirus and a greater proportion of them used antimicrobials before coming to the hospital.     

Mechanical and In Vitro Biological Performance of Graphene Nanoplatelets Reinforced Calcium Silicate Composite

Calcium silicate (CaSiO₃, CS) ceramic composites reinforced with graphene nanoplatelets (GNP) were prepared using hot isostatic pressing (HIP) at 1150°C. Quantitative microstructural analysis suggests that GNP play a role in grain size and is responsible for the improved densification. Raman spectroscopy and scanning electron microscopy showed that GNP survived the harsh processing conditions of the selected HIP processing parameters. The uniform distribution of 1 wt.% GNP in the CS matrix, high densification and fine CS grain size help to improve the fracture toughness by ∼130%, hardness by ∼30% and brittleness index by ∼40% as compared to the CS matrix without GNP. The toughening mechanisms, such as crack bridging, pull-out, branching and deflection induced by GNP are observed and discussed. The GNP/CS composites exhibit good apatite-forming ability in the simulated body fluid (SBF). Our results indicate that the addition of GNP decreased pH value in SBF. Effect of addition of GNP on early adhesion and proliferation of human osteoblast cells (hFOB) was measured in vitro. The GNP/CS composites showed good biocompatibility and promoted cell viability and cell proliferation. The results indicated that the cell viability and proliferation are affected by time and concentration of GNP in the CS matrix.