Nucleoporins mediate nucleocytoplasmic exchange of macromolecules and several have been assigned active mitotic functions. Nucleoporins can participate in various mitotic functions like spindle assembly, kinetochore organisation and chromosome segregation- important for genome integrity. Pathways to genome integrity are frequently deregulated in cancer and many are regulated in part by microRNAs. Indeed, altered levels of numerous microRNAs have frequently been associated with tumorigenesis. Here, we unveil a microRNA-mediated regulation of the nucleoporin Nup214 and its downstream effect on genome integrity.
Databases/bioinformatic tools such as miRBase, Oncomine and RNAhybrid predicted Nup214 as a miR-133b target. To validate this, we used luciferase reporter assays, Real-Time PCR and immuno-blotting. Flow cytometry and immuno-blots of mitotic markers were used to analyse cell cycle pattern upon thymidine synchronization and miR-133b treatment. Mitotic indices and chromosomal abnormalities were assessed by immuno-fluorescence for FITC-tagged phospho-H3 as well as video-microscopy for GFP-tagged histone H4. Annexin V/propidium iodide staining, caspase3/PARP cleavage and colony formation assays were done to investigate cell death upon either miR-133b transfection or NUP214 knockdown by siRNA. UPCI:SCC084, HCT116, HeLa-H4-pEGFP and HEK293 (human oral squamous cell carcinoma, colorectal, cervical carcinomas and embryonic kidney cell lines, respectively) were used. miR-133b and NUP214 expressions were validated in cancer cell lines and tissues by Real-Time PCR.
Examination of head and neck tumour tissues and cancer cell lines revealed that Nup214 and miR-133b expressions are negatively correlated. In vitro, Nup214 was significantly downregulated by ectopic miR-133b. This downregulation elevated mitotic indices and delayed degradation of mitotic marker proteins cyclinB1 and cyclinA and dephosphorylation of H3. Moreover, this mitotic delay enhanced chromosomal abnormalities and apoptosis.
We have identified NUP214, a member of the massive nuclear pore complex, as a novel miR-133b target. Thus, we have shown a hitherto unknown microRNA regulation of mitosis mediated by a member of the nucleoporin family. Based on observations, we also raise some hypotheses regarding transport-dependent/independent functions of Nup214 in this study. Our results hence attempt to explain why miR-133b is generally downregulated in tumours and lay out the potential for Nup214 as a therapeutic target in the treatment of cancer.
Electronic supplementary material
The online version of this article (doi:10.1186/s12943-015-0299-z) contains supplementary material, which is available to authorized users.
Nucleoporin; Nup214; miR-133b; Mitosis; Apoptosis; Cell cycle; MicroRNA; Cancer; Head and neck cancer; Chromosomal abnormality
Radicicol, an antifungal antibiotic, was previously identified as a compound having antimalarial activity. However, its mechanism of action in Plasmodium falciparum was not elucidated. While characterizing its antimalarial function, we observed that radicicol manifested two distinct developmental defects in cultured P. falciparum in a concentration-dependent manner. At a low concentration of radicicol, a significant percentage of drug-treated parasites were arrested at the schizont stage, while at a higher concentration, the parasites were unable to multiply from schizont to ring. Also, the newly formed rings and trophozoites were extremely delayed in development, eventually leading to cell death. We intended to characterize the potential molecular target of radicicol at its sublethal doses. Our results demonstrated that radicicol specifically impaired mitochondrial replication. This decrement was associated with a severalfold increment of the topoisomerase VIB transcript as well as protein in treated cells over that of untreated parasites. Topoisomerase VIB was found to be localized in the organelle fraction. Our docking study revealed that radicicol fits into the Bergerat fold of Pf topoisomerase VIB present in its ATPase domain. Altogether, these data allow us to conclude that P. falciparum topoisomerase VIB might be one of the targets of radicicol causing inhibition of mitochondrial replication. Hence, radicicol can be suitably employed to explore the mitochondrial physiology of malaria parasites.
Urinary tract infection (UTI) is one of the most common infectious diseases at the community level. In order to assess the adequacy of empirical therapy, the susceptibility of antibiotics and resistance pattern of bacteria responsible for UTI in West Bengal, India, were evaluated throughout the period of 2008–2013. The infection reports belonging to all age groups and both sexes were considered. Escherichia coli was the most abundant uropathogen with a prevalence rate of 67.1%, followed by Klebsiella spp. (22%) and Pseudomonas spp. (6%). Penicillin was least effective against UTI-causing E. coli and maximum susceptibility was recorded for the drugs belonging to fourth generation cephalosporins. Other abundant uropathogens, Klebsiella spp., were maximally resistant to broad-spectrum penicillin, followed by aminoglycosides and third generation cephalosporin. The antibiotic resistance pattern of two principal UTI pathogens, E. coli and Klebsiella spp. in West Bengal, appears in general to be similar to that found in other parts of the Globe. Higher than 50% resistance were observed for broad-spectrum penicillin. Fourth generation cephalosporin and macrolides seems to be the choice of drug in treating UTIs in Eastern India. Furthermore, improved maintenance of infection incident logs is needed in Eastern Indian hospitals in order to facilitate regular surveillance of the occurrence of antibiotic resistance patterns, since such levels continue to change.
antibiotic susceptibility; uropathogens; West Bengal; India; pathogenic bacteria
Prostate gland is a fibromusculoglandular structure situated at the neck of urinary bladder. So, enlargement or growth of prostate due to nodular hyperplasia (NHP) or prostatic intraepithelial neoplasia (PIN) or adenocarcinoma may give rise to bladder outlet obstruction. Malignant growth i.e., PIN or adenocarcinoma cases are associated with increased blood level of prostate-specific antigen (PSA) and increased expression of different sex-steroid receptors because the growth is dependent on the interactions of androgen, progesterone and estrogen. The aim of our study is to correlate the histopathology, PSA levels and expression of different sex-steroid receptors by immunohistochemistry in different prostatic growth lesions. Among the total 50 cases received, inclusive of transurethral resection of prostate (TURP), transrectal ultrasound-guided biopsy and radical prostatectomy, 34 cases were diagnosed as NHP, 4 cases as PIN and 12 cases as adenocarcinoma histopathologically. Serum PSA values above 10 ng/ml were seen in 2 cases of PIN and 11 cases of adenocarcinoma and none of NHP. Estrogen receptor (ER) () expressions were negative in all cases. Progesterone receptor (PR) expressions were strongly positive in 35% cases of both NHP and adenocarcinoma, whereas androgen receptor (AR) expressions were strong among all cases of adenocarcinoma and only in four cases of NHP. By observing these findings it can be suggested that antiandrogen and antiprogesterone therapy simultaneously will do better than antiandrogen alone in treating prostatic growth lesions.
Androgen receptor; immunohistochemistry; progesterone receptor; prostatic growth; prostate-specific antigen
Thyroid disorders are common in pregnancy and the most common disorder is subclinical hypothyroidism. Due to the complex hormonal changes during pregnancy, it is important to remember that thyroxine requirements are higher in pregnancy. According to recent American Thyroid Association (ATA) guidelines, the recommended reference ranges for TSH are 0.1 to 2.5 mIU/L in the first trimester, 0.2 to 3.0 mIU/L in the second trimester, and 0.3 to 3.0 mIU/L in the third trimester. Maternal hypothyroidism is an easily treatable condition that has been associated with increased risk of low birth weight, fetal distress, and impaired neuropsychological development. Hyperthyroidism in pregnancy is less common as conception is a problem. Majority of them are due to Graves’ disease, though gestational hyperthyroidism is to be excluded. Preferred drug is propylthiouracil (PTU) with the target to maintain free T4 in upper normal range. Doses can be reduced in third trimester due to the immune-suppressant effects of pregnancy. Early and effective treatment of thyroid disorder ensures a safe pregnancy with minimal maternal and neonatal complications.
Thyroid disorders; pregnancy; hypothyroidism
Sundarbans is the single largest deltaic mangrove forest in the world, formed at estuarine phase of the Ganges - Brahmaputra river system. Primary productivity of marine and coastal phytoplankton contributes to 15% of global oceanic production. But unfortunately estuarine dynamics of tropical and subtropical estuaries have not yet received proper attention in spite of the fact that they experience considerable anthropogenic interventions and a baseline data is required for any future comparison. This study is an endeavor to this end to estimate the primary productivity (gross and net), community respiration and nitrification rates in different rivers and tidal creeks around Jharkhali island, a part of Sundarbans estuary surrounded by the mangrove forest during a period of three years starting from November’08 to October’11.
Various physical and chemical parameters of water column like pH, temperature, conductivity, dissolved oxygen, turbidity, suspended particulate matter, secchi disc index, tidal fluctuation and tidal current velocity, standing crop and nutrients were measured along with water column productivity. Relationship of net water column productivity with algal biomass (standing crop), nutrient loading and turbidity were determined experimentally. Correlations of bacterial abundance with community respiration and nitrification rates were also explored. Annual integrated phytoplankton production rate of this tidal estuary was estimated to be 151.07 gC m-2 y-1. Gross primary productivity showed marked inter annual variation being lowest in monsoon and highest in postmonsoon period.
Average primary production was a function of nutrient loading and light penetration in the water column. High aquatic turbidity, conductivity and suspended particulate matter were the limiting factors to attenuate light penetration with negative influence on primary production. Community respiration and nitrification rates of the estuary were influenced by the bacterial abundance. The estuary was phosphorus limited in postmonsoon whereas nitrogen-limited in premonsoon and monsoon period. High algal biomass and primary productivity indicated the estuary to be in eutrophic state in most of the time throughout the year. Our study also indicated a seasonal shifting between autotrophic and heterotrophic conditions in Sundarban estuarine ecosystem and it is a tropical, well mixed (high tidal influx) and marine dominated (no fresh water connection) system.
Net ecosystem metabolism; Gross primary productivity; Community respiration; Nitrification; Nutrient load; Sundarban estuary
Repairing double strand breaks (DSBs) is absolutely essential for the survival of obligate intracellular parasite Toxoplasma gondii. Thus, DSB repair mechanisms could be excellent targets for chemotherapeutic interventions. Recent genetic and bioinformatics analyses confirm the presence of both homologous recombination (HR) as well as non homologous end joining (NHEJ) proteins in this lower eukaryote. In order to get mechanistic insights into the HR mediated DSB repair pathway in this parasite, we have characterized the key protein involved in homologous recombination, namely TgRad51, at the biochemical and genetic levels. We have purified recombinant TgRad51 protein to 99% homogeneity and have characterized it biochemically. The ATP hydrolysis activity of TgRad51 shows a higher KM and much lower kcat compared to bacterial RecA or Rad51 from other related protozoan parasites. Taking yeast as a surrogate model system we have shown that TgRad51 is less efficient in gene conversion mechanism. Further, we have found that TgRad51 mediated gene integration is more prone towards random genetic loci rather than targeted locus. We hypothesize that compromised ATPase activity of TgRad51 is responsible for inefficient gene targeting and poor gene conversion efficiency in this protozoan parasite. With increase in homologous flanking regions almost three fold increments in targeted gene integration is observed, which is similar to the trend found with ScRad51. Our findings not only help us in understanding the reason behind inefficient gene targeting in T. gondii but also could be exploited to facilitate high throughput knockout as well as epitope tagging of Toxoplasma genes.
In recent years, Hsp90 is found to interact with several telomeric proteins at various phases of cell cycle. The Hsp90 chaperone system controls assembly and disassembly of telomere structures and thus maintains the dynamic state of telomere. Here, for the first time we report that the activity of another telomeric protein Sir2p is modulated by Hsp82, the ortholog of Hsp90 from budding yeast (Saccharomyces cerevisiae). In a temperature sensitive Hsp90 deficient yeast strain (iG170Dhsp82), less abundant Sir2p is observed, resulting in de-repression of telomere silencing and a complete loss of mating type silencing. Intriguingly, over expression of Hsp90, either by exposing cells to heat shock or by introducing HSP82 overexpression plasmid also yields reduced level of Sir2p, with a consequential loss of telomere silencing. Thus, Hsp90 homeostasis maintains the cellular pool of Sir2p and thereby controls the reversible nature of telomere silencing. Interestingly, such regulation is independent of one of its major co-chaperones Sba1 (human ortholog of p23).
Sundarbans is the largest chunk of mangrove forest and only tiger mangrove land in the world. Compared to the rich species diversity and uniqueness, very few studies have so far been conducted here, mainly due to its inaccessibility. This study explores water quality, density of biomass, species diversity, phytoplankton abundance and bacterial population of a tidal creek in Sunderban estuary during the post and pre monsoon period of 2008-09.
Phytoplankton community was observed to be dominated by diatoms (Biacillariophyceae) followed by Pyrrophyceae (Dinoflagellates) and Chlorophyceae. A total of 46 taxa belonging to 6 groups were recorded. Other algal groups were Cyanophyceae, Euglenophyceae and Chrysophyceae. Species diversity was highest in summer (March) and lowest in winter season (November) in all the sample stations indicating its close correlation with ambient temperature. Species evenness was fairly high in all five stations throughout the study period. Present study indicated that dissolved oxygen, nutrients and turbidity are the limiting factors for the phytoplankton biomass. The estuary was in eutrophic condition (Chlorophyll-a ≥10 μg/L) in winter. During the month of May phytoplankton biomass declined and at high salinity level (21.2PSU) new phytoplankton species take over, which are definitely better resilient to the high saline environment. Bio-indicator species like Polykrikos schwartzil, Dinophysis norvegica and Prorocentrum concavum points to moderately polluted water quality of the estuary.
Eutrophication as well as presence of toxic Dinoflagellates and Cyanophyceae in the tidal creek of Sundarban estuary definitely revealed the deteriorated status of the water quality. The structure and function of the mangrove food web is unique, driven by both marine and terrestrial components. But little attention has been paid so far to the adaptive responses of mangrove biota to the various disturbances, and now our work unfolds the fact that marine status of Sundarban estuary is highly threatened which in turn will affect the ecology of the mangrove. This study indicates that ecosystem dynamics of the world heritage site Sundarban may facilitate bioinvasion putting a question mark on the sustainability of mangroves.
The minor physical anomaly (MPA) is believed to reflect abnormal development of the CNS. The aim is to find incidence of MPA and its behavioral correlates in Down syndrome and to compare these findings with the other causes of intellectual disability and normal population.
Materials and Methods:
One-hundred and forty intellectually disabled people attending a tertiary care set-up and from various NGOs are included in the study. The age-matched group from normal population was also studied for comparison. MPA are assessed by using Modified Waldrop scale and behavioral abnormality by Diagnostic assessment scale for severely handicapped (DASH II scale).
The Down syndrome group had significantly more MPA than other two groups and most of the MPA is situated in the global head region. There is strong correlation (P < 0.001) between the various grouped items of Modified Waldrop scale. Depression subscale is correlated with anomalies in the hands (P < 0.001), feet and Waldrop total items (P < 0.005). Mania item of DASH II scale is related with anomalies around the eyes (P < 0.001). Self-injurious behavior and total Waldrop score is negatively correlated with global head.
Down syndrome group has significantly more MPA and a pattern of correlation between MPA and behavioral abnormalities exists which necessitates a large-scale study.
Behavioral abnormalities; correlation; Down syndrome; minor physical anomaly
Agenesis of corpus callosum can have various neuropsychiatric manifestations. Following case report highlights the case of a young man presenting with features of recurrent brief depressive disorder, each lasting for about 3 to 7 days, for over a year. He had history of occasional headache and episodes of swooning attack in between, usually precipitated by emotional events. His neuroimaging revealed agenesis of corpus callosum. He was experiencing swooning attacks as he became aware that some ‘unusual’ findings were present in his reports. Recurrent brief depression can be a manifestation of this congenital anomaly, and conversion disorder can be present as comorbid diagnosis perhaps due to ignorance and fear of this apparently innocuous congenital malformation.
Conversion disorder; corpus callosum agenesis; recurrent depression
The protein kinase C (PKC) family regulates macrophage function involved in host defense against infection. In the case of Leishmania donovani infection, the impairment of PKC-mediated signaling is one of the crucial events for the establishment of parasite into the macrophages. Earlier reports established that C-C chemokines mediated protection against leishmaniasis via the generation of nitric oxide after 48 h. In this study, we investigated the role of MIP-1α and MCP-1 in the regulation of impaired PKC activity in the early hours (6 h) of infection. These chemokines restored Ca2+-dependent PKC activity and inhibited Ca2+-independent atypical PKC activity in L. donovani-infected macrophages under both in vivo and in vitro conditions. Pretreatment of macrophages with chemokines induced superoxide anion generation by activating NADPH oxidase components in infected cells. Chemokine administration in vitro induced the migration of infected macrophages and triggered the production of reactive oxygen species. In vivo treatment with chemokines significantly restricted the parasitic burden in livers as well as in spleens. Collectively, these results indicate a novel regulatory role of C-C chemokines in controlling the intracellular growth and multiplication of L. donovani, thereby demonstrating the antileishmanial properties of C-C chemokines in the disease process.
Type 2 diabetes is an emerging health challenge all over the world as a result of urbanization, high prevalence of obesity, sedentary lifestyle and other stress related factors compounded with the genetic prevalence. The health consequences and economic burden of the obesity and related diabetes mellitus epidemic are enormous. Different signaling molecules secreted by adipocytes have been implicated in the development of obesity and associated insulin resistance in type 2 diabetes. Human adiponectin, a 244-amino acid collagen-like protein is solely secreted by adipocytes and acts as a hormone with anti-inflammatory and insulin-sensitizing properties. Adiponectin secretion, in contrast to secretion of other adipokines, is paradoxically decreased in obesity which may be attributable to inhibition of adiponectin gene transcription. There are several mechanisms through which adiponectin may decrease the risk of type 2 diabetes, including suppression of hepatic gluconeogenesis, stimulation of fatty acid oxidation in the liver, stimulation of fatty acid oxidation and glucose uptake in skeletal muscle, and stimulation of insulin secretion. To date, no systematic review has been conducted that evaluate the potential importance of adiponectin metabolism in insulin resistance. In this review attempt has been made to explore the relevance of adiponectin metabolism for the development of diabetes mellitus. This article also identifies this novel target for prospective therapeutic research aiming successful management of diabetes mellitus.
Adiponectin; Obesity; Dyslipidemia; Type 2 diabetes mellitus; Insulin resistance
Plasmodium falciparum causes the most severe form of malaria and affects 3.2 million people annually. Due to the increasing incidence of resistance to existing drugs, there is a growing need to discover new and more effective drugs against malaria. Despite the global importance of P. falciparum, vast majority of its proteins are uncharacterized experimentally. Application of newer approaches using several “omics” data has become successful for exploring the biological interactions underlying cellular processes. Till date not many system level study has been published using P. falciparum protein protein interaction. Hence, the purpose of this study is to develop a standardized pipeline for structural, functional, and topographical analysis of large scale protein protein interaction network (PPIN) in order to identify proteins important for network topology and integrity. Here, P. falciparum PPIN has been utilized as a model for better understanding of the molecular mechanisms of survival and pathogenesis of malaria parasite.
Various graph theoretical approaches were implemented to identify highly interacting hub and central proteins that are crucial for network integrity. Further, potential network perturbing proteins via an in-silico knock-out (KO) analysis to isolate important interacting proteins (IIPs), which in principle, can elicit significant impact on the global and local environments of the P. falciparum interaction network.
177 hubs and 132 central proteins were identified from the malarial (proteins: 1607; interactions: 4750) PPI networks. Using the in-silico knock-out exercise 131 and 99 global and local network perturbing proteins were also identified. Finally, 271 proteins from P. falciparum were shortlisted as important interacting proteins (IIPs), which not only play crucial role in intra-pathogen network integrity, stage specificity but also interact with various human proteins involved in multiple metabolic pathways within the host cell. These IIPs could be used as potential drug targets in malarial research.
Graph theoretical analysis of PPIN can be a very useful approach to identify proteins that are important for regulation of the interactions required for an organism’s survival. Important interacting proteins (IIPs) identified using P. falciparum PPIN provides a useful dataset containing probable candidates for future drug target analysis.
Electronic supplementary material
The online version of this article (doi:10.1186/s12936-015-0562-1) contains supplementary material, which is available to authorized users.
Centrality analysis; IIPs; Plasmodium; Graph theory; Host pathogen interaction; Hubs
Visceral leishmaniasis (VL), which is endemic as a major infectious disease in the tropical and subtropical countries, is caused by a protozoan parasite Leishmania donovani. At present, restricted treatment options and lack of vaccines intensify the problem of controlling VL. Therefore, finding a novel immunoprophylactic or therapeutic principle is a pressing need. Here, we report that arabinosylated lipoarabinomannan (Ara-LAM), a TLR2-ligand isolated from Mycobacterium smegmatis, exhibits a strong immunomodulatory property that conferred protection against L. donovani infection. Although, Ara-LAM modulates TLR2 and MAPK signaling, it is not known whether Ara-LAM involves IFN-γ signaling for effective parasite clearance. Because, it is reported that IFN-γ signaling, a principle mediator of NO generation and macrophage and Tcell activation, is hampered during leishmanial pathogenesis. Ara-LAM increases IFN-γ receptor expression and potentiates IFN-γ receptor signaling through JAK-STAT pathway. Moreover, Ara-LAM reciprocally modulates IRF4 and IRF8 expression and reinstates anti-leishmanial Th1 response that eventuates in significantly reduced parasite load in spleen and liver of L. donovani-infected BALB/c mice. IFN-γRα silencing resulted in the suppression of these host-protective mechanisms affected by Ara-LAM. Thus, Ara-LAM-mediated restoration of IFN-γ responsiveness is a novel immuno-modulatory principle for protection against L. donovani susceptible host.
We evaluate the clinical performance of the Luminex xTAG gastrointestinal (GI) pathogen in vitro diagnostic (IVD) assay in a comparison between clinical and public health laboratories. The site reproducibility study showed 98.7% sensitivity with high positive and negative agreement values (96.2% and 99.8%, respectively), while assay performance against confirmatory methods resulted in 96.4% sensitivity with similar positive and negative agreement values (90.1% and 99.5%, respectively). High-throughput detection of multiple GI pathogens improved turnaround time, consolidated laboratory workflow, and simplified stool culture practices, thus reducing the overall cost and number of specimens processed.
In utero ethanol exposure causes Fetal Alcohol Spectrum Disorders, associated with reduced brain plasticity; the mechanisms of these effects are not well understood, particularly with respect to glial involvement.
Astrocytes release factors that modulate neurite outgrowth. We explored the hypothesis that ethanol inhibits neurite outgrowth by increasing the release of inhibitory chondroitin sulfate proteoglycans (CSPGs) from astrocytes.
Astrocyte treatment with ethanol inhibited the activity of arylsulfatase B (ARSB), the enzyme that removes sulfate groups from chondroitin-4-sulfate (C4S) and triggers the degradation of C4S, increased total sulfated glycosaminoglycans (GAGs), C4S, and neurocan core-protein content and inhibited neurite outgrowth in neurons co-cultured with ethanol-treated astrocytes in vitro, effects reversed by treatment with recombinant ARSB.
Ethanol also inhibited ARSB activity and increased sulfate GAG and neurocan levels in the developing hippocampus after in vivo ethanol exposure.
ARSB silencing increased the levels of sulfated GAGs, C4S, and neurocan in astrocytes and inhibited neurite outgrowth in co-cultured neurons, indicating that ARSB activity directly regulates C4S and affects neurocan expression.
In summary, this study reports two major findings: ARSB modulates sulfated GAG and neurocan levels in astrocytes and astrocyte-mediated neurite outgrowth in co-cultured neurons; and ethanol inhibits the activity of ARSB, increases sulfated GAG, C4S, and neurocan levels, and thereby inhibits astrocyte-mediated neurite outgrowth.
An unscheduled increase in CSPGs in the developing brain may lead to altered brain connectivity and to premature decrease in neuronal plasticity and therefore represents a novel mechanism by which ethanol can exert its neurodevelopmental effects.
Astrocytes; arylsulfatase B; fetal alcohol; chondroitin sulfate; neurocan
ensure high fidelity in translation, many aminoacyl-tRNA synthetases,
enzymes responsible for attaching specific amino acids to cognate
tRNAs, require proof-reading mechanisms. Most bacterial prolyl-tRNA
synthetases (ProRSs) misactivate alanine and employ a post-transfer
editing mechanism to hydrolyze Ala-tRNAPro. This reaction
occurs in a second catalytic site (INS) that is distinct from the
synthetic active site. The 2′-OH of misacylated tRNAPro and several conserved residues in the Escherichia coli ProRS INS domain are directly involved in Ala-tRNAPro deacylation. Although mutation of the strictly conserved lysine
279 (K279) results in nearly complete loss of post-transfer editing
activity, this residue does not directly participate in Ala-tRNAPro hydrolysis. We hypothesized that the role of K279 is to
bind the phosphate backbone of the acceptor stem of misacylated tRNAPro and position it in the editing active site. To test this
hypothesis, we carried out pKa, charge
neutralization, and free-energy of binding calculations. Site-directed
mutagenesis and kinetic studies were performed to verify the computational
results. The calculations revealed a considerably higher pKa of K279 compared to an isolated lysine and
showed that the protonated state of K279 is stabilized by the neighboring
acidic residue. However, substitution of this acidic residue with
a positively charged residue leads to a significant increase in Ala-tRNAPro hydrolysis, suggesting that enhancement in positive charge
density in the vicinity of K279 favors tRNA binding. A charge-swapping
experiment and free energy of binding calculations support the conclusion
that the positive charge at position 279 is absolutely necessary for
tRNA binding in the editing active site.
Functionalized polymer materials with ion exchange groups and integration of nano-structured materials is an emerging area for catalytic and water pollution control applications. The polymerization of materials such as acrylic acid often requires persulfate initiator and a high temperature start. However, is generally known that metal ions accelerate such polymerizations starting from room temperature. If the metal is properly selected, it can be used in environmental applications adding two advantages simultaneously. This paper deals with this by polymerizing acrylic acid using iron as accelerant and its subsequent use for nanoparticle synthesis in hydrogel and PVDF membranes. Characterizations of hydrogel, membranes and nanoparticles were carried out with different techniques. Nanoparticles sizes of 30–60 nm were synthesized. Permeability and swelling measurements demonstrate an inverse relationship between hydrogel mesh size (6.30 to 8.34 nm) and membrane pores (222 to 110 nm). Quantitative reduction of trichloroethylene/chloride generation by Fe/Pd nanoparticles in hydrogel/membrane platforms was also performed.
TCE; xerogel; iron nanoparticles; membrane functionalization; responsive behavior; catalysis
Pathologically elevated serum levels of fibroblast growth factor-23 (FGF23), a bone-derived hormone that regulates phosphorus homeostasis, result in renal phosphate wasting and lead to rickets or osteomalacia. Rarely, elevated serum FGF23 levels are found in association with mosaic cutaneous disorders that affect large proportions of the skin and appear in patterns corresponding to the migration of ectodermal progenitors. The cause and source of elevated serum FGF23 is unknown. In those conditions, such as epidermal and large congenital melanocytic nevi, skin lesions are variably associated with other abnormalities in the eye, brain and vasculature. The wide distribution of involved tissues and the appearance of multiple segmental skin and bone lesions suggest that these conditions result from early embryonic somatic mutations. We report five such cases with elevated serum FGF23 and bone lesions, four with large epidermal nevi and one with a giant congenital melanocytic nevus. Exome sequencing of blood and affected skin tissue identified somatic activating mutations of HRAS or NRAS in each case without recurrent secondary mutation, and we further found that the same mutation is present in dysplastic bone. Our finding of somatic activating RAS mutation in bone, the endogenous source of FGF23, provides the first evidence that elevated serum FGF23 levels, hypophosphatemia and osteomalacia are associated with pathologic Ras activation and may provide insight in the heretofore limited understanding of the regulation of FGF23.
Rationale: Treatment of pulmonary nontuberculous mycobacteria, especially Mycobacterium abscessus, requires prolonged, multidrug regimens with high toxicity and suboptimal efficacy. Options for refractory disease are limited.
Objectives: We reviewed the efficacy and toxicity of inhaled amikacin in patients with treatment-refractory nontuberculous mycobacterial lung disease.
Methods: Records were queried to identify patients who had inhaled amikacin added to failing regimens. Lower airway microbiology, symptoms, and computed tomography scan changes were assessed together with reported toxicity.
Measurements and Main Results: The majority (80%) of the 20 patients who met entry criteria were women; all had bronchiectasis, two had cystic fibrosis and one had primary ciliary dyskinesia. At initiation of inhaled amikacin, 15 were culture positive for M. abscessus and 5 for Mycobacterium avium complex and had received a median (range) of 60 (6, 190) months of mycobacterial treatment. Patients were followed for a median of 19 (1, 50) months. Eight (40%) patients had at least one negative culture and 5 (25%) had persistently negative cultures. A decrease in smear quantity was noted in 9 of 20 (45%) and in mycobacterial culture growth for 10 of 19 (53%). Symptom scores improved in nine (45%), were unchanged in seven (35%), and worsened in four (20%). Improvement on computed tomography scans was noted in 6 (30%), unchanged in 3 (15%), and worsened in 11 (55%). Seven (35%) stopped amikacin due to: ototoxicity in two (10%), hemoptysis in two (10%), and nephrotoxicity, persistent dysphonia, and vertigo in one each.
Conclusions: In some patients with treatment-refractory pulmonary nontuberculous mycobacterial disease, the addition of inhaled amikacin was associated with microbiologic and/or symptomatic improvement; however, toxicity was common. Prospective evaluation of inhaled amikacin for mycobacterial disease is warranted.
amikacin; nontuberculous mycobacteria; therapeutics; antibacterial agents
Familial Alzheimer's Disease (FAD) mutations in presenilin (PS) modulate PS/γ-secretase activity and therefore contribute to AD pathogenesis. Previously, we found that PS/γ-secretase cleaves voltage-gated sodium channel β2 subunits (Navβ2), releases the intracellular domain of Navβ2 (β2-ICD), and thereby increases intracellular sodium channel α-subunit Nav1.1 levels. Here, we tested whether FAD-linked PS1 mutations modulate Navβ2 cleavages and Nav1.1 levels.
To analyze the effects of PS1-linked FAD mutations on Navβ2 processing and Nav1.1 levels in neuronal cells.
We first generated B104 rat neuroblastoma cells stably expressing Navβ2 and wild-type PS1, PS1 with one of three FAD mutations (E280A, M146L, or Δ-E9), or PS1 with a non-FAD mutation (D333G). Navβ2 processing and Nav1.1 protein and mRNA levels were then analyzed by Western blot and real-time RT-PCR, respectively.
The FAD-linked E280A mutation significantly decreased PS/γ-secretase-mediated processing of Navβ2 as compared to wild-type PS1 controls, both in cells and a cell-free system. Nav1.1 mRNA and protein levels, as well as the surface levels of Nav channel α-subunits, were also significantly reduced in PS1 (E280A) cells.
Our data indicate that the FAD-linked PS1 E280A mutation decreases Nav channel levels by partially inhibiting PS/γ-secretase-mediated cleavage of Navβ2 in neuronal cells.
Alzheimer's disease; presenilin; voltage-gated sodium channel
Lung lesions may develop from tissue reactions to known or unknown stimuli and present with different morphological descriptions. The pathogenesis may be induced and maintained by different bioactive substances, of which, the upregulation matrix metalloproteinases (MMPs) play a vital role. Inhibition of the MMPs, therefore, may be a prospective mode of therapy for such lesions.
Materials and Methods:
A number of patients with lung lesions of different morphologies and presentations were treated empirically with long-term oral doxycycline (100 mg BID) upon exclusion of malignancy and infection in an open, single-arm, prospective, observational pilot study. The effect of the treatment was recorded on serial x-rays/computed tomography (CT) scans and the impact of treatment was measured with a visual analog scale (VAS) or a Likert-like scale. Furthermore, six independent pulmonologists’ opinion (expressed on a ‘0’ to ‘100’ scale) were pooled with regard to the significance and the expectedness of such a change.
Twenty-six patients (mean age 49.33 years and male: female ratio = 10:3) with different types of pulmonary parenchymal/pleural lesions were treated with long-term oral doxycycline for a mean duration of 386.88 days related to the available radiological comparison. They showed a mean improvement of 3.99 on the Likert-like scale and 78% on the VAS scale. The mean significance of the change was 83.33%, with a mean expectedness of 18% as per the pooled opinion of the pulmonologists.
The significant and unexpected resolution of different tissue lesions from long-term doxycycline appears to be a novel observation. This needs proper scientific validation.
Doxycycline; lung lesions; matrix metalloproteinase