Exploring the diversity and metabolic potential of actinomycetes from temperate marine sediments from Newfoundland, Canada.

Marine sediments from Newfoundland, Canada were explored for biotechnologically promising Actinobacteria using culture-independent and culture-dependent approaches. Culture-independent pyrosequencing analyses uncovered significant actinobacterial diversity (H'-2.45 to 3.76), although the taxonomic diversity of biotechnologically important actinomycetes could not be fully elucidated due to limited sampling depth. Assessment of culturable actinomycete diversity resulted in the isolation of 360 actinomycetes representing 59 operational taxonomic units, the majority of which (94 %) were Streptomyces. The biotechnological potential of actinomycetes from NL sediments was assessed by bioactivity and metabolomics-based screening of 32 representative isolates. Bioactivity was exhibited by 41 % of isolates, while 11 % exhibited unique chemical signatures in metabolomics screening. Chemical analysis of two isolates resulted in the isolation of the cytotoxic metabolite 1-isopentadecanoyl-3ß-D-glucopyranosyl-X-glycerol from Actinoalloteichus sp. 2L868 and sungsanpin from Streptomyces sp. 8LB7. These results demonstrate the potential for the discovery of novel bioactive metabolites from actinomycetes isolated from Atlantic Canadian marine sediments.

Assessment of diabetic macrovascular complications: a prediabetes model.

Prediabetes is a condition that requires early intervention against diabetic macrovascular complications. This study aims to assess whether or not the likelihood of diabetes macrovascular complications occurring in prediabetes can be better estimated by a model combining a set of conventional and emerging biomarkers, with a view to improving cardiovascular disease (CVD) screening in individuals with elevated blood glucose levels associated with prediabetes. A total of 71 participants (female/male: 32/39) were divided into two groups - the prediabetic group (preDM: n=34) and the diabetic with cardiovascular complications group (DM+CVD: n=37). Blood glucose level (BGL), blood pressure (BP), total cholesterol (TC), high-density lipoprotein (HDL) and TC:HDL ratio, erythrocyte oxidative stress (as determined by reduced glutathione [GSH], malondialdehyde and methaemoglobin levels) and vascular events (D-dimer, homocysteine and whole blood viscosity) were measured. Statistical analysis was by binomial logistic regression modelling with forward likelihood ratio step procedures. A combination of BGL, BP, erythrocyte GSH and TC gave the best group identifications, with 28/34 (82.4%) and 29/37 (78.4%) members correctly identified in the preDM and DM + CVD groups, respectively. Six of the 34 (17.6%) prediabetes individuals were logistically identified as having diabetic macrovascular complications, but clinically did not qualify for CVD intervention under current screening models. The authors propose that a combination of BGL, BP, erythrocyte GSH and TC can provide a clinically acceptable standard for identifying CVD risk in individuals with prediabetes. This model provides a tool for early identification and targeted intervention in individuals with subclinical diabetes who are at risk of CVD.

Microfluidic and cross-linking methods for encapsulation of living cells and bacteria - A review.

Microencapsulation of living cells is a field that has been heavily investigated by many researchers over the past two decades. Numerous experimental setups have been developed to encapsulate living cells in microbeads using different microfluidic devices and materials. Previous studies have investigated different microfluidic devices and materials for use in cancer treatment, drug delivery, environmental remediation, food production, and cell culture contexts. Some of the current challenges to these setups are maintaining reasonable levels of cell viability, cell leaching, nutrient and oxygen diffusion, and ensuring uniform microbead shape and size distribution. Addressing these issues and identifying the most reproducible and convenient setup enables researchers to efficiently encapsulate living cells and further advance the biomedical field. The efficiency of microencapsulation in terms of cell viability and uniform microbead shape and size distribution are directly related to the type of device used and the cross-linking method applied. Hence, the focus of this review is to assess the effects of using T-junction, flow-focusing, and co-flow microfluidic devices as well as thermal, ionic, and photo cross-linking methods for the microencapsulation of living cells. Recent applications of bacteria microencapsulation using microfluidic systems since 2017 are presented.

Oxidative damage indices for the assessment of subclinical diabetic macrovascular complications.

Subclinical cardiovascular disease (SCVD), including complications in diabetes, is associated with oxidative damage and precedes future cardiovascular disease (CVD). Hence, assessment and management of oxidative damage is imperative. This study investigates biomarkers associated with CVD, diabetes and oxidative stress in order to determine a set of indices that could be useful to assess oxidative damage in diabetic macrovascular pathogenesis. A total of 266 participants were selected and divided into seven groups (control, family history of diabetes, prediabetes, prediabetes with CVD, diabetes mellitus [DM], DM+CVD and CVD) based on clinical history/status. Blood glucose (BG) level, erythrocyte glutathione (GSH), malondialdehyde, methaemoglobin, D-dimer, homocysteine, blood viscosity and cholesterol profile were determined. Factorial MANOVA and independent univariate analyses were performed. Prevalence of significant biomarkers was assessed following a 3.5-year retrospective study. Multivariate analysis showed statistically significant differences between groups (P < 0.0001) with post hoc tests identifying a statistically significant association for BG level (P < 0.0001), GSH (P < 0.0001), D-dimer (P < 0.02) and total cholesterol (P < 0.0001). Of the subjects who showed hyperglycaemia-associated progression in clinical and biochemistry status, 89% had low-level GSH and 44% had high-level D-dimer. Four individuals exhibited prediabetic status at some stage and would qualify for macrovascular disease intervention. The results of this study suggest that BG level, D-dimer, GSH and total cholesterol contribute significantly to a diabetic oxidative damage panel of markers that could assist in evidence-based pharmacological intervention with anti-aggregation and/or antioxidant agents against future CVD in diabetes.

Temperature and pH effect on glucose production from pretreated bagasse by a novel species of Citrobacter and other bacteria.

Cellulolytic bacteria that produce cellulases, which are active over a range of pH and temperatures, can be used to catalyze hydrolysis of pretreated lignocellulosic material. This is important in the production of second generation biofuels among other biotechnological applications. In this investigation, bacteria isolated from sugarcane bagasse were identified as strains of Enterobacter xiangfangensis, Serratia rubidaea, Klebsiella pneumoniae and a novel species of Citrobacter designated Citrobacter sp. UWIBGS10. The glucose production potential of these strains was studied on thermally and solvent pretreated sugarcane bagasse. This was performed at 24-hour intervals up to 168 hours in the range of pH 5-9 and temperature range 25-40 °C. Maximal concentrations of glucose for Citrobacter sp. UWIBGS10 occurred at pH 6 and 25 °C. For E. xiangfangensis, S. rubidaea, K. pneumoniae glucose concentrations were consistent across the pH and temperature ranges examined. From these results it could be concluded that the bacteria demonstrated ability for lignocellulolytic hydrolysis for the production of glucose and could be further explored for the characterization of commercial cellulolytic enzymes.

Preclinical pharmacology of the natural product anticancer agent bryostatin 1, an activator of protein kinase C.

Bryostatin 1, a natural product anticancer agent isolated from a marine bryozoan, has been shown in tissue culture to activate protein kinase C. This agent has recently undergone Phase I testing in humans given either as a bolus i.v. injection or a continuous infusion. To understand how bryostatin 1 might be used best as an anticancer agent, a study of the pharmacokinetics, tissue distribution, metabolism, and elimination of bryostatin 1 in mice was undertaken, using [C26-3H]-labeled bryostatin 1. Following i.v. administration, the plasma disappearance curve for bryostatin 1 could be described by a two-compartment model, with half-lives of 1.05 and 22.97 h, respectively. In contrast, the plasma disappearance curve for bryostatin 1 administered i.p. was better described by a first order absorption one-compartment model, with an absorption half-life of 0.81 h and an elimination half-life of 28.76 h, respectively. The majority of radioactivity in plasma was associated with the intact drug for up to 24 h after dosing. In the first 12 h after i.v administration, urinary excretion represented the major pathway of elimination, with 23.0 +/- 1.9% (mean +/- SD) of the administered dose excreted. Within 72 h after i.v. administration, approximately equal amounts of radioactivity (40%) were excreted in feces compared to urine. Bryostatin 1 was widely distributed in many organs but concentrated in the lung, liver, gastrointestinal tract, and fatty tissue. The concentration in the gastrointestinal tract, along with the fecal excretion, suggests the possibility of enterohepatic circulation of this drug. In summary, this study demonstrates that bryostatin 1 is relatively stable in vivo, widely distributed but concentrated in some major tissues, and rapidly excreted first through urine and at later times through the feces. The data from this animal study should be useful in the design of future human trials with this anticancer drug.

In vivo and in vitro investigations into the biosynthetic relatedness of the pseudopterosins.

[formula: see text] Both in vivo and in vitro techniques have been developed to test putative intermediates in the biosynthetic pathway to the pseudopterosins, antiinflammatory compounds isolated from Pseudopterogorgia elisabethae. Furthermore, specific activity data we have obtained indicate that pseudopterosin A is a precursor to pseudopterosins B, C, and D. We conclude that in the biosynthesis xylose is attached to the diterpene skeleton to produce pseudopterosin A and is then aceylated to form pseudopterosins B-D.

Three distinct neuronal phenotypes exist in embryonic rat hippocampal neurons cultured in basic fibroblast growth factor.

The possibility that neurons cultured in basic fibroblast growth factor (bFGF) are heterogeneous raises concerns about their subsequent use in gene transfection and transplantation studies. We have examined the fate of embryonic hippocampal neurons in bFGF culture, and now conclude that these cells are not only heterogeneous, but possess neurons of various stages of development. Morphological and immunocytochemical analysis reveal three distinct developmental classes of neurons are present in extended bFGF culture. This tripartite classification is supported by electrophysiological analysis, which reveals that upon depolarization, neurons with precursor and juvenile neuron morphologies are unable to fire action potentials. The third class of neurons, which resemble age-matched polarized neurons in standard serum culture, fired multiple action potentials indicative of a mature phenotype. These data show neurons at multiple developmental stages co-exist in bFGF culture, and provide an experimental basis for their classification.

Long-term bFGF neuronal culture: reintroduction into serum medium yields neurons and non-neuronal cells with neuronal characteristics.

The potential use of bFGF immortalized cells as hosts for delivering foreign genes into nervous tissue led us to examine the effect of maintaining, E-18 hippocampal neurons for extended periods in bFGF culture prior to transfer into a standard, serum-containing, medium. We found: (1) many, if not most, precursors seen in bFGF, mature into glia and not into primary neurons after medium exchange; (2) the electrophysiology of the neurons which do mature after medium transfer and replating, is similar to that of neurons in standard cultures; (3) extended culture in bFGF prior to cell harvesting and replating into standard medium generates neurons from the precursors that possess proper neuronal polarization, morphology, and electrophysiology; and (4) extended bFGF also induces the expression, on transfer into standard medium, of an additional cell type with a distinct non-neuronal morphology that stains with the neuronal marker MAP-2. These results illustrate the need for additional characterization of long-term growth factor effects on maintained progenitor cells prior to their use in gene therapy and transplantation.

Purification and characterization of the fatty acid synthase from Bugula neritina.

The fatty acid synthase from Bugula neritina has been purified 100-fold using ammonium sulfate precipitation, ion-exchange and size exclusion chromatography. The purified enzyme has a molecular weight of approximately 382,000 Da, as judged by gel filtration. Polyacrylamide gel electrophoresis under denaturing conditions in the presence of SDS revealed one major protein band of approximately 190,000 Da suggesting that the enzyme is a homodimer. The size of the enzyme, together with the observation that the FAS activity is independent of the concentration of acyl carrier protein, indicate that the FAS from Bugula neritina is a type I. A detailed analysis of the products of the purified FAS indicated that palmitic acid is the primary product and longer chain fatty acids are not produced.

The effects of postoperative continuous passive motion on peripheral nerve repair and regeneration. An experimental investigation in rabbits.

The effects of continuous passive motion (CPM) on nerve regeneration following nerve repair were investigated. In 26 rabbits, the medial popliteal nerve was transected and microsurgically repaired. Half of the animals were treated with cast immobilization and the rest with 70 degrees arc CPM. Both treatments were discontinued on day 14. After sacrifice on day 100, no animal showed separation at the suture line. Mean nerve conduction velocity was slightly slower in the CPM than in the immobilization group. Mean fibre density was also slightly less in the CPM group but the difference was not significant. Mean fibre diameters, fibre diameter distributions, and soleus-muscle wet weights were similar in the two groups.

Osteoarthritis. A primary care approach for physicians in 2000 and beyond.

Osteoarthritis is one of the most common disorders presented to the primary care physician in the over 50 years age group in the Kingdom of Saudi Arabia. The diagnosis is made by history, typical x-ray findings and non-contributory laboratory investigations. The understanding of the pathogenesis of the condition is undergoing change. The development of osteoarthritis is dependent on age, sex, genetic predisposition, and previous trauma to the joint and abnormal mechanical forces caused primarily by obesity. Biochemically, there is an imbalance in the enzymes of cartilage degradation and cartilage regeneration. Management in 2000 focuses on patient education, appropriate exercise, relief of pain through judicious combination of capsaicin cream, acetaminophen in appropriate dose, and nonsteroidal anti-inflammatory drug therapy. The latter is undergoing revolutionary change with the introduction of the Coxiella-2 specific inhibitors, Rofecoxib and Celecoxib in the autumn of 2000 to the Kingdom. With these agents the primary care physician has an effective analgesic therapy, once a day dosing and a dramatic reduction in nonsteroidal anti-inflammatory drug gastropathy across all groups of patients. Finally, when the conservative management by the primary care physician is of benefit no longer, judicious referral to an experienced Orthopedic Surgeon for the modern surgical approaches should be given.

In vivo and in vitro biosynthesis of saponins in sea cucumbers.

The triterpene precursor of saponins in sea cucumbers has been identified as parkeol [lanost-9(11)-en-3 beta-ol] [1]. Dissection of the sea cucumbers Holothuria floridea and Actinopyga agassize after incubations with radiolabeled parkeol demonstrated that saponin biosynthesis occurs exclusively in the Cuvier gland. This result was corroborated by incubating a cell-free extract of the Cuvier gland with labeled parkeol and observing transformation of the precursor to saponins.

Effects of continuous passive motion and immobilization on synovitis and cartilage degradation in antigen induced arthritis.

OBJECTIVE: To determine the effects of continuous passive motion (CPM) and immobilization on synovitis and cartilage degradation in an experimental model of chronic inflammatory, antigen-induced arthritis. METHODS: After bilateral arthritis induction of knee joints in 22 NZW rabbits, one knee was immobilized with a flexion splint while the opposite knee received CPM. RESULTS: After 2 weeks (n = 10), the CPM treated knees had significantly greater joint swelling, synovial effusion, and histologic synovitis scores compared to its opposite immobilized knees. However, the total cartilage degradation score showed no statistically significant difference between the two treatments. When the treatments were discontinued after 2 weeks and animals were allowed intermittent active motion of both knees in cages for 4 weeks (n = 12), no statistically significant difference in joint swelling, synovial effusion, and histologic synovitis score was observed between the 2 treatments. The articular cartilage degradation, however, was significantly greater in the immobilized knees compared to its opposite CPM treated knees. Five of 12 immobilized knees had articular surface erosion compared to none in the CPM treated knees. Loss of cellularity was also significantly greater in the immobilized knees. CONCLUSION: Although CPM produced greater synovitis at 2 weeks, articular cartilage was better preserved in the knees treated with CPM than immobilization at 6 weeks.