OBJECTIVE

To report the refractive, topographic, and clinical outcomes 3 years after corneal collagen cross-linking (CXL) in eyes with progressive keratoconus.

METHODS

Prospective, randomized controlled trial.

METHODS

One hundred eyes with progressive keratoconus were randomized into the CXL treatment or control groups.

METHODS

Cross-linking was performed by instilling riboflavin 0.1% solution containing 20% dextran for 15 minutes before and during the 30 minutes of ultraviolet A irradiation (3 mW/cm(2)). Follow-up examinations were arranged at 3, 6, 12, 24, and 36 months.

METHODS

The primary outcome measure was the maximum simulated keratometry value (Kmax). Other outcome measures were uncorrected visual acuity (UCVA; measured in logarithm of the minimum angle of resolution [logMAR] units), best spectacle-corrected visual acuity (BSCVA; measured in logMAR units), sphere and cylinder on subjective refraction, spherical equivalent, minimum simulated keratometry value, corneal thickness at the thinnest point, endothelial cell density, and intraocular pressure.

RESULTS

The results from 48 control and 46 treated eyes are reported. In control eyes, Kmax increased by a mean of 1.20±0.28 diopters (D), 1.70±0.36 D, and 1.75±0.38 D at 12, 24, and 36 months, respectively (all P <0.001). In treated eyes, Kmax flattened by -0.72±0.15 D, -0.96±0.16 D, and -1.03±0.19 D at 12, 24, and 36 months, respectively (all P <0.001). The mean change in UCVA in the control group was +0.10±0.04 logMAR (P = 0.034) at 36 months. In the treatment group, both UCVA (-0.15±0.06 logMAR; P = 0.009) and BSCVA (-0.09±0.03 logMAR; P = 0.006) improved at 36 months. There was a significant reduction in corneal thickness measured using computerized videokeratography in both groups at 36 months (control group: -17.01±3.63 μm, P <0.001; treatment group: -19.52±5.06 μm, P <0.001) that was not observed in the treatment group using the manual pachymeter (treatment group: +5.86±4.30 μm, P = 0.181). The manifest cylinder increased by 1.17±0.49 D (P = 0.020) in the control group at 36 months. There were 2 eyes with minor complications that did not affect the final visual acuity.

CONCLUSIONS

At 36 months, there was a sustained improvement in Kmax, UCVA, and BSCVA after CXL, whereas eyes in the control group demonstrated further progression.

Mucosal-associated invariant T (MAIT) cells express an invariant T cell receptor (TCR) α-chain (TRAV1-2 joined to TRAJ33, TRAJ20, or TRAJ12 in humans), which pairs with an array of TCR β-chains. MAIT TCRs can bind folate- and riboflavin-based metabolites restricted by the major histocompatibility complex (MHC)-related class I-like molecule, MR1. However, the impact of MAIT TCR and MR1-ligand heterogeneity on MAIT cell biology is unclear. We show how a previously uncharacterized MR1 ligand, acetyl-6-formylpterin (Ac-6-FP), markedly stabilized MR1, potently up-regulated MR1 cell surface expression, and inhibited MAIT cell activation. These enhanced properties of Ac-6-FP were attributable to structural alterations in MR1 that subsequently affected MAIT TCR recognition via conformational changes within the complementarity-determining region (CDR) 3β loop. Analysis of seven TRBV6-1(+) MAIT TCRs demonstrated how CDR3β hypervariability impacted on MAIT TCR recognition by altering TCR flexibility and contacts with MR1 and the Ag itself. Ternary structures of TRBV6-1, TRBV6-4, and TRBV20(+) MAIT TCRs in complex with MR1 bound to a potent riboflavin-based antigen (Ag) showed how variations in TRBV gene usage exclusively impacted on MR1 contacts within a consensus MAIT TCR-MR1 footprint. Moreover, differential TRAJ gene usage was readily accommodated within a conserved MAIT TCR-MR1-Ag docking mode. Collectively, MAIT TCR heterogeneity can fine-tune MR1 recognition in an Ag-dependent manner, thereby modulating MAIT cell recognition.

OBJECTIVE

Chronic heart failure (CHF) is a common and leading cause of death in industrialized countries. The potential benefits of micronutrient supplementation in CHF are extensive. Therefore, we examined the influence of long-term multiple micronutrient supplementation on left ventricular (LV) function, levels of pro-inflammatory cytokines, and quality-of-life (QoL) in elderly patients with CHF.

RESULTS

Thirty CHF patients [age 75.4 (0.7), mean (SEM), LV ejection fraction (LVEF) < or =35%] were randomized to receive capsules containing a combination of high-dose micronutrients (calcium, magnesium, zinc, copper, selenium, vitamin A, thiamine, riboflavin, vitamin B(6), folate, vitamin B(12), vitamin C, vitamin E, vitamin D, and Coenzyme Q10) or placebo for 9 months in a double-blind fashion. All subjects were on stable optimal medical therapy for at least 3 months before enrolment. At randomization and at study end, tumour necrosis factor-alpha and its soluble receptors TNFR-1 and TNFR-2 were measured and six-minute walk test and QoL were assessed. Cardiac magnetic resonance scanning was performed to evaluate cardiac dimensions and LVEF. Two patients died during follow-up. The remaining patients (14 randomized to placebo and 14 to micronutrients) were well matched for LV function, symptoms, and exercise capacity. At the end of the follow-up period, LV volumes were reduced in the intervention group with no change in the placebo group [-13.1 (17.1)% vs. +3.8 (10.0)%; P<0.05]. LVEF increased by 5.3+/-1.4% in the intervention group and was unchanged in the placebo group (P<0.05). Patients taking micronutrients also had a significant improvement in QoL score between enrolment and study end [+9.5 (1.6)%; P<0.05], whereas those taking placebo had a slight deterioration [-1.1 (0.8)%; P=0.12]. Six-minute walk test and inflammatory cytokine levels remained unchanged in both groups.

CONCLUSIONS

Long-term multiple micronutrient supplementation can improve LV volumes and LVEF and QoL scores in elderly patients with heart failure due to LV systolic dysfunction.

Defining the essential genome of bacterial pathogens is central to developing an understanding of the biological processes controlling disease. This has proven elusive for Pseudomonas aeruginosa during chronic infection of the cystic fibrosis (CF) lung. In this paper, using a Monte Carlo simulation-based method to analyze high-throughput transposon sequencing data, we establish the P. aeruginosa essential genome with statistical precision in laboratory media and CF sputum. Reconstruction of the global requirements for growth in CF sputum compared with defined growth conditions shows that the latter requires several cofactors including biotin, riboflavin, and pantothenate. Comparison of P. aeruginosa strains PAO1 and PA14 demonstrates that essential genes are primarily restricted to the core genome; however, some orthologous genes in these strains exhibit differential essentiality. These results indicate that genes with similar molecular functions may have distinct genetic roles in different P. aeruginosa strains during growth in CF sputum. We also show that growth in a defined growth medium developed to mimic CF sputum yielded virtually identical fitness requirements to CF sputum, providing support for this medium as a relevant in vitro model for CF microbiology studies.

Cancer stem cells (CSCs) are thought to drive tumor growth, metastasis and chemoresistance. Although surface markers such as CD133 and CD44 have been successfully used to isolate CSCs, their expression is not exclusively linked to the CSC phenotype and is prone to environmental alteration. We identified cells with an autofluorescent subcellular compartment that exclusively showed CSC features across different human tumor types. Primary tumor-derived autofluorescent cells did not overlap with side-population (SP) cells, were enriched in sphere culture and during chemotherapy, strongly expressed pluripotency-associated genes, were highly metastatic and showed long-term in vivo tumorigenicity, even at the single-cell level. Autofluorescence was due to riboflavin accumulation in membrane-bounded cytoplasmic structures bearing ATP-dependent ABCG2 transporters. In summary, we identified and characterized an intrinsic autofluorescent phenotype in CSCs of diverse epithelial cancers and used this marker to isolate and characterize these cells.

Sea cucumbers, belonging to the class Holothuroidea, are marine invertebrates, habitually found in the benthic areas and deep seas across the world. They have high commercial value coupled with increasing global production and trade. Sea cucumbers, informally named as bêche-de-mer, or gamat, have long been used for food and folk medicine in the communities of Asia and Middle East. Nutritionally, sea cucumbers have an impressive profile of valuable nutrients such as Vitamin A, Vitamin B1 (thiamine), Vitamin B2 (riboflavin), Vitamin B3 (niacin), and minerals, especially calcium, magnesium, iron and zinc. A number of unique biological and pharmacological activities including anti-angiogenic, anticancer, anticoagulant, anti-hypertension, anti-inflammatory, antimicrobial, antioxidant, antithrombotic, antitumor and wound healing have been ascribed to various species of sea cucumbers. Therapeutic properties and medicinal benefits of sea cucumbers can be linked to the presence of a wide array of bioactives especially triterpene glycosides (saponins), chondroitin sulfates, glycosaminoglycan (GAGs), sulfated polysaccharides, sterols (glycosides and sulfates), phenolics, cerberosides, lectins, peptides, glycoprotein, glycosphingolipids and essential fatty acids. This review is mainly designed to cover the high-value components and bioactives as well as the multiple biological and therapeutic properties of sea cucumbers with regard to exploring their potential uses for functional foods and nutraceuticals.

The genome sequencing of Buchnera aphidicola BCc from the aphid Cinara cedri, which is the smallest known Buchnera genome, revealed that this bacterium had lost its symbiotic role, as it was not able to synthesize tryptophan and riboflavin. Moreover, the biosynthesis of tryptophan is shared with the endosymbiont Serratia symbiotica SCc, which coexists with B. aphidicola in this aphid. The whole-genome sequencing of S. symbiotica SCc reveals an endosymbiont in a stage of genome reduction that is closer to an obligate endosymbiont, such as B. aphidicola from Acyrthosiphon pisum, than to another S. symbiotica, which is a facultative endosymbiont in this aphid, and presents much less gene decay. The comparison between both S. symbiotica enables us to propose an evolutionary scenario of the transition from facultative to obligate endosymbiont. Metabolic inferences of B. aphidicola BCc and S. symbiotica SCc reveal that most of the functions carried out by B. aphidicola in A. pisum are now either conserved in B. aphidicola BCc or taken over by S. symbiotica. In addition, there are several cases of metabolic complementation giving functional stability to the whole consortium and evolutionary preservation of the actors involved.

Cobalamin in the form of adenosylcobalamin (Ado-CBL) is known to repress expression of genes for vitamin B(12) biosynthesis and be transported by a posttranscriptional regulatory mechanism, which involves direct binding of Ado-CBL to 5'untranslated gene regions (5'UTR). Using comparative analysis of genes and regulatory regions, we identified a highly conserved RNA structure, the B12-element, which is widely distributed in 5'UTRs of vitamin B(12)-related genes in eubacteria. Multiple alignment of approximately 200 B12-elements from 66 bacterial genomes reveals their common secondary structure and several extended regions of sequence conservation, including the previously known B12-box motif. In analogy to the model of regulation of the riboflavin and thiamin biosynthesis, we suggest Ado-CBL-mediated regulation based on formation of alternative RNA structures including the B12-element. In Gram-negative proteobacteria, as well as in cyanobacteria, actinobacteria, and the CFB group, the cobalamin biosynthesis and vitamin B(12) transport genes are predicted to be regulated by inhibition of translation initiation, whereas in the Bacillus/Clostridium group of Gram-positive bacteria, these genes seem to be regulated by transcriptional antitermination. Phylogenetic analysis of the B12-elements reveals a large number of likely duplications of B12-elements in several bacterial genomes. These lineage-specific duplications of RNA regulatory elements seem to be a major evolutionary mechanism for expansion of the vitamin B(12) regulon.

Our knowledge of the mechanisms and regulation of intestinal absorption of water-soluble vitamins under normal physiological conditions, and of the factors/conditions that affect and interfere with theses processes has been significantly expanded in recent years as a result of the availability of a host of valuable molecular/cellular tools. Although structurally and functionally unrelated, the water-soluble vitamins share the feature of being essential for normal cellular functions, growth and development, and that their deficiency leads to a variety of clinical abnormalities that range from anaemia to growth retardation and neurological disorders. Humans cannot synthesize water-soluble vitamins (with the exception of some endogenous synthesis of niacin) and must obtain these micronutrients from exogenous sources. Thus body homoeostasis of these micronutrients depends on their normal absorption in the intestine. Interference with absorption, which occurs in a variety of conditions (e.g. congenital defects in the digestive or absorptive system, intestinal disease/resection, drug interaction and chronic alcohol use), leads to the development of deficiency (and sub-optimal status) and results in clinical abnormalities. It is well established now that intestinal absorption of the water-soluble vitamins ascorbate, biotin, folate, niacin, pantothenic acid, pyridoxine, riboflavin and thiamin is via specific carrier-mediated processes. These processes are regulated by a variety of factors and conditions, and the regulation involves transcriptional and/or post-transcriptional mechanisms. Also well recognized now is the fact that the large intestine possesses specific and efficient uptake systems to absorb a number of water-soluble vitamins that are synthesized by the normal microflora. This source may contribute to total body vitamin nutrition, and especially towards the cellular nutrition and health of the local colonocytes. The present review aims to outline our current understanding of the mechanisms involved in intestinal absorption of water-soluble vitamins, their regulation, the cell biology of the carriers involved and the factors that negatively affect these absorptive events.

OBJECTIVE

Collagen crosslinking of the cornea has been developed recently as a quasiconservative treatment of keratoconus. Biomechanical in vitro measurements have demonstrated a significant increase in biomechanical stiffness of the crosslinked cornea. The aim of the present study was to evaluate the effect of this new procedure on the collagen fiber diameter of the rabbit cornea.

METHODS

The corneas of the right eyes of 10 New Zealand White albino rabbits were crosslinked by application of the photosensitizer riboflavin and exposure to UVA light (370 nm, 3 mW/cm2) for 30 minutes. The left fellow control eyes were either left untreated (rabbits 1-4), deepithelialized (rabbits 5-7), or deepithelialized and treated with riboflavin/dextran solution (rabbits 8-10) to exclude an influence of epithelial debridement or hydration changes on the fiber diameter. On ultrathin sections of samples from the anterior and posterior cornea, the collagen fiber diameter was measured semiautomatically with the help of morphometric computer software.

RESULTS

In the anterior stroma, the collagen fiber diameter in the treated corneas was significantly increased by 12.2% (3.96 nm), and in the posterior stroma by 4.6% (1.63 nm), compared with the control fellow eyes. In the crosslinked eyes, the collagen fiber diameter was also significantly increased by, on average, 9.3% (3.1 nm) in the anterior compared with the posterior stroma within the same eye.

CONCLUSIONS

Collagen crosslinking using riboflavin and UVA leads to a significant increase in corneal collagen diameter. This alteration is the morphologic correlate of the crosslinking process leading to an increase in biomechanical stability. The crosslinking effect is strongest in the anterior half of the stroma because of the rapid decrease in UVA irradiance across the corneal stroma as a result of riboflavin-enhanced UVA absorption.

OBJECTIVE

Corneal collagen cross-linking by UVA/riboflavin (X-linking) represents a new method for the treatment of progressive keratoconus and currently is under clinical study. To avoid UVA irradiation damage to the corneal endothelium, the parameters for X-linking are set in a way that effective treatment occurs only in the first 300 microm of the corneal stroma. Here, X-linking not only strengthens the biomechanical properties of the cornea but also induces keratocyte apoptosis. To date, the effectiveness of treatment could be monitored only indirectly by postoperative follow-up corneal topographies or using corneal confocal microscopy. Here we describe a corneal stromal demarcation line indicating the transition zone between cross-linked anterior corneal stroma and untreated posterior corneal stroma. The demarcation line is biomicroscopically detectable in slit-lamp examination as early as 2 weeks after treatment.

METHODS

X-linking was performed in 16 cases of progressive keratoconus, and corneas were examined biomicroscopically and by means of corneal topography and pachymetry before and after treatment.

RESULTS

In 14 of 16 cases, a thin stromal demarcation line was visible at a depth of approximately 300 microm over the whole cornea after X-linking treatment.

CONCLUSIONS

This newly observed demarcation line may result from differences in the refractive index and/or reflection properties of untreated versus X-linked corneal stroma and represents an effective tool to biomicroscopically easily monitor the depth of effective X-linking treatment in keratoconus.

OBJECTIVE

To assess ultrastructural stromal modifications after riboflavin-UVA-induced cross-linking of corneal collagen in patients with progressive keratoconus.

METHODS

This was a second-phase prospective nonrandomized open study in 10 patients with progressive keratoconus treated by riboflavin-UVA-induced cross-linking of corneal collagen and assessed by means of Heidelberg Retinal Tomograph II Rostock Corneal Module (HRT II-RCM) in vivo confocal microscopy. The eye in the worst clinical condition was treated for each patient. Treatment under topical anesthesia included corneal deepithelization (9-mm diameter) and instillation of 0.1% riboflavin phosphate-20% dextran T 500 solution at 5 minutes before UVA irradiation and every 5 minutes for a total of 30 minutes. UVA irradiation was 7 mm in diameter. Patients were assessed by HRT II-RCM confocal microscopy in vivo at 1, 3, and 6 months after treatment.

RESULTS

Rarefaction of keratocytes in the anterior and intermediate stroma, associated with stromal edema, was observed immediately after treatment. The observation at 3 months after the operation detected keratocyte repopulation in the central treated area, whereas the edema had disappeared. Cell density increased progressively over the postoperative period. At approximately 6 months, keratocyte repopulation was complete, accompanied by increased density of stromal fibers. No endothelial damage was observed at any time.

CONCLUSIONS

Reduction in anterior and intermediate stromal keratocytes followed by gradual repopulation has been confirmed directly in vivo in humans by HRT II-RCM confocal microscopy after riboflavin-UVA-induced corneal collagen cross-linking.

Mucosal-associated invariant T (MAIT) cells express a semi-invariant T cell receptor (TCR) that detects microbial metabolites presented by the nonpolymorphic major histocompatibility complex (MHC)-like molecule MR1. The highly conserved nature of MR1 in conjunction with biased MAIT TCRα chain usage is widely thought to indicate limited ligand presentation and discrimination within a pattern-like recognition system. Here, we evaluated the TCR repertoire of MAIT cells responsive to three classes of microbes. Substantial diversity and heterogeneity were apparent across the functional MAIT cell repertoire as a whole, especially for TCRβ chain sequences. Moreover, different pathogen-specific responses were characterized by distinct TCR usage, both between and within individuals, suggesting that MAIT cell adaptation was a direct consequence of exposure to various exogenous MR1-restricted epitopes. In line with this interpretation, MAIT cell clones with distinct TCRs responded differentially to a riboflavin metabolite. These results suggest that MAIT cells can discriminate between pathogen-derived ligands in a clonotype-dependent manner, providing a basis for adaptive memory via recruitment of specific repertoires shaped by microbial exposure.

The Mtr respiratory pathway of Shewanella oneidensis strain MR-1 is required to effectively respire both soluble and insoluble forms of oxidized iron. Flavins (riboflavin and flavin mononucleotide) recently have been shown to be excreted by MR-1 and facilitate the reduction of insoluble substrates. Other Shewanella species tested accumulated flavins in supernatants to an extent similar to that of MR-1, suggesting that flavin secretion is a general trait of the species. External flavins have been proposed to act as both a soluble electron shuttle and a metal chelator; however, at biologically relevant concentrations, our results suggest that external flavins primarily act as electron shuttles for MR-1. Using deletion mutants lacking various Mtr-associated proteins, we demonstrate that the Mtr extracellular respiratory pathway is essential for the reduction of flavins and that decaheme cytochromes found on the outer surface of the cell (MtrC and OmcA) are required for the majority of this activity. Given the involvement of external flavins in the reduction of electrodes, we monitored current production by Mtr respiratory pathway mutants in three-electrode bioreactors under controlled flavin concentrations. While mutants lacking MtrC were able to reduce flavins at 50% of the rate of the wild type in cell suspension assays, these strains were unable to grow into productive electrode-reducing biofilms. The analysis of mutants lacking OmcA suggests a role for this protein in both electron transfer to electrodes and attachment to surfaces. The parallel phenotypes of Mtr mutants in flavin and electrode reduction blur the distinction between direct contact and the redox shuttling strategies of insoluble substrate reduction by MR-1.

The purpose of this review is to summarize the most important human clinical trials of antioxidants as cancer prevention agents conducted to date, provide an overview of currently ongoing studies, and discuss future steps needed to advance research in this field. To date there have been several large (at least 7000 participants) trials testing the efficacy of antioxidant supplements in preventing cancer. The specific agents (diet-derived direct antioxidants and essential components of antioxidant enzymes) tested in those trials included β-carotene, vitamin E, vitamin C, selenium, retinol, zinc, riboflavin, and molybdenum. None of the completed trials produced convincing evidence to justify the use of traditional antioxidant-related vitamins or minerals for cancer prevention. Our search of ongoing trials identified six projects at various stages of completion. Five of those six trials use selenium as the intervention of interest delivered either alone or in combination with other agents. The lack of success to date can be explained by a variety of factors that need to be considered in the next generation research. These factors include lack of good biological rationale for selecting specific agents of interest; limited number of agents tested to date; use of pharmacological, rather than dietary, doses; and insufficient duration of intervention and follow-up. The latter consideration underscores the need for alternative endpoints that are associated with increased risk of neoplasia (i.e., biomarkers of risk), but are detectable prior to tumor occurrence. Although dietary antioxidants are a large and diverse group of compounds, only a small proportion of candidate agents have been tested. In summary, the strategy of focusing on large high-budget studies using cancer incidence as the endpoint and testing a relatively limited number of antioxidant agents has been largely unsuccessful. This lack of success in previous trials should not preclude us from seeking novel ways of preventing cancer by modulating oxidative balance. On the contrary, the well demonstrated mechanistic link between excessive oxidative stress and carcinogenesis underscores the need for new studies. It appears that future large-scale projects should be preceded by smaller, shorter, less expensive biomarker-based studies that can serve as a link from mechanistic and observational research to human cancer prevention trials. These relatively inexpensive studies would provide human experimental evidence for the likely efficacy, optimum dose, and long-term safety of the intervention of interest that would then guide the design of safe, more definitive large-scale trials.

OBJECTIVE

Combined riboflavin/UVA treatment inducing collagen cross-links in the cornea has been shown to increase the biomechanical rigidity of the cornea and has been used successfully in the treatment of progressive keratoconus. The current study was undertaken to investigate the possible cytotoxic effect of combined riboflavin/UVA treatment on corneal keratocytes in vivo.

METHODS

Thirty-four New Zealand white rabbits were treated with 0.1% riboflavin solution and surface UVA irradiances ranging from 0.75 to 4 mW/cm2 (1.35- 7.2 J/cm2) for 30 minutes. The animals were euthanized either 4 (n = 6) or 24 (n = 28) hours postoperatively. Four additional control eyes underwent epithelial debridement alone. The corneas of the enucleated eyes were evaluated in routine histologic sections. In addition, the TUNEL technique and transmission electron microscopy were used for the detection of keratocyte apoptosis.

RESULTS

In the control eyes with corneal epithelial debridement only, apoptotic keratocytes were found in the anterior 50 microm of the corneal stroma 4 hours postoperatively. However, riboflavin/UVA-induced apoptosis was only visible in the rabbit eyes enucleated 24 hours postoperatively. In these eyes, we found apoptosis of keratocytes down to a variable stromal depth depending on the applied UVA irradiance. A cytotoxic UVA irradiance for keratocytes in the range of 0.5-0.7 mW/cm2 could be deduced.

CONCLUSIONS

Riboflavin/UVA treatment leads to a dose-dependent keratocyte damage that can be expected in human corneas down to a depth of 300 microm using a surface UVA dose of 5.4 J/cm2. Future studies should be done to examine the keratocyte repopulation and exclude possible adverse sequelae of keratocyte loss like stromal scarring or thinning.

OBJECTIVE

To describe which dietary nutrient variables are related to subjective and objective habitual sleep and subjective and objective napping.

METHODS

Participants were 459 post-menopausal women enrolled in the Women's Health Initiative. Objective sleep was estimated using one week of actigraphy. Subjective sleep was prospectively estimated with a daily sleep diary. Dietary nutrients were calculated from food frequency questionnaires.

RESULTS

The most significant correlations were with subjective napping, including (from strongest to weakest): total fat, calories, saturated fat, monounsaturated fat, trans fat, water, proline, serine, tyrosine, phenylalanine, valine, cholesterol, leucine, glutamic acid, ash, isoleucine, histidine, sodium, tryptophan, protein, threonine, cystine, methionine, phosphorous, polyunsaturated fat, animal protein, aspartic acid, arginine, lysine, alanine, caffeine, riboflavin, gamma-tocopherol, glycine, retinol, delta-tocopherol, Vitamin D, and selenium. Actigraphic nocturnal sleep duration was negatively associated with total fat, monounsaturated fat, trans fat, saturated fat, polyunsaturated fat, calories, gamma-tocopherol, cholesterol, and alpha-tocopherol-eq.

CONCLUSIONS

Actigraphic total sleep time was negatively associated with intake of fats. Subjective napping, which may be a proxy for subjective sleepiness, was significantly related to fat intake as well as intake of meat.

BACKGROUND

Bariatric surgery is widely performed to induce weight loss.

OBJECTIVE

The objective was to examine changes in vitamin status after 2 bariatric surgical techniques.

METHODS

A randomized controlled trial was conducted in 2 Scandinavian hospitals. The subjects were 60 superobese patients [body mass index (BMI; in kg/m(2)): 50-60]. The surgical interventions were either laparoscopic Roux-en-Y gastric bypass or laparoscopic biliopancreatic diversion with duodenal switch. All patients received multivitamins, iron, calcium, and vitamin D supplements. Gastric bypass patients also received a vitamin B-12 substitute. The patients were examined before surgery and 6 wk, 6 mo, and 1 y after surgery.

RESULTS

Of 60 surgically treated patients, 59 completed the follow-up. After surgery, duodenal switch patients had lower mean vitamin A and 25-hydroxyvitamin D concentrations and a steeper decline in thiamine concentrations than did the gastric bypass patients. Other vitamins (riboflavin, vitamin B-6, vitamin C, and vitamin E adjusted for serum lipids) did not change differently in the surgical groups, and concentrations were either stable or increased. Furthermore, duodenal switch patients had lower hemoglobin and total cholesterol concentrations and a lower BMI (mean reduction: 41% compared with 30%) than did gastric bypass patients 1 y after surgery. Additional dietary supplement use was more frequent among duodenal switch patients (55%) than among gastric bypass patients (26%).

CONCLUSIONS

Compared with gastric bypass, duodenal switch may be associated with a greater risk of vitamin A and D deficiencies in the first year after surgery and of thiamine deficiency in the initial months after surgery. Patients who undergo these 2 surgical interventions may require different monitoring and supplementation regimens in the first year after surgery. This trial was registered at ClinicalTrials.gov as NCT00327912.

Chickpea (Cicer arietinum L.) is an important pulse crop grown and consumed all over the world, especially in the Afro-Asian countries. It is a good source of carbohydrates and protein, and protein quality is considered to be better than other pulses. Chickpea has significant amounts of all the essential amino acids except sulphur-containing amino acids, which can be complemented by adding cereals to the daily diet. Starch is the major storage carbohydrate followed by dietary fibre, oligosaccharides and simple sugars such as glucose and sucrose. Although lipids are present in low amounts, chickpea is rich in nutritionally important unsaturated fatty acids such as linoleic and oleic acids. β-Sitosterol, campesterol and stigmasterol are important sterols present in chickpea oil. Ca, Mg, P and, especially, K are also present in chickpea seeds. Chickpea is a good source of important vitamins such as riboflavin, niacin, thiamin, folate and the vitamin A precursor β-carotene. As with other pulses, chickpea seeds also contain anti-nutritional factors which can be reduced or eliminated by different cooking techniques. Chickpea has several potential health benefits, and, in combination with other pulses and cereals, it could have beneficial effects on some of the important human diseases such as CVD, type 2 diabetes, digestive diseases and some cancers. Overall, chickpea is an important pulse crop with a diverse array of potential nutritional and health benefits.

OBJECTIVE

To evaluate the possible cytotoxic effect of combined riboflavin-ultraviolet-A (UVA) treatment on the corneal endothelium.

METHODS

Department of Ophthalmology, Technical University of Dresden, Dresden, Germany.

METHODS

The right eyes of 34 New Zealand White rabbits were treated with riboflavin and various endothelial UVA doses ranging from 0.16 to 0.9 J/cm2 (0.09 to 0.5 mW/cm2, 370 nm) and postoperative enucleation times of 4 hours and 24 hours. The endothelial cells were evaluated in histological sections. The terminal deoxynulceotidyl transferase deoxy-UTP-nick-end labeling (TUNEL) technique and transmission electron microscopy were used to detect apoptosis.

RESULTS

There was no endothelial damage in the 6 rabbit eyes enucleated at 4 hours. In those enucleated at 24 hours, there was significant necrosis and apoptosis of endothelial cells in the corneas treated with an endothelial dose of>> or =0.65 J/cm2 (0.36 mW/cm2), which is about twice the endothelial UVA dose used in the treatment of keratoconus patients.

CONCLUSIONS

In rabbit corneas with a corneal thickness less than 400 microm, the endothelial UVA dose reached a cytotoxic level of>> or =0.65 J/cm2 (0.36 mW/cm2) using the standard surface UVA dose of 5.4 J/cm2 (3 mW/cm2). Pachymetry should be routinely performed before riboflavin-UVA treatment; in thinner corneas, irradiation should not be done because of the cytotoxic risk to the endothelium.

OBJECTIVE

To test the biomechanical efficiency of corneal crosslinking with riboflavin without epithelial debridement (C3-R).

METHODS

Moscow Helmholtz Research Institute of Eye Diseases, Moscow, Russia.

METHODS

The left eyes of rabbits were crosslinked using standard crosslinking including epithelial removal (Group 1), using benzalkonium chloride-containing proxymetacaine eyedrops without epithelial removal (Group 2), or using preservative-free oxybuprocaine eyedrops without epithelial removal (Group 3). All left eyes received riboflavin solution and were irradiated with an ultraviolet-A double diode for 30 minutes (irradiance 3 mW/cm(2)). The animals were killed 1 day after crosslinking. Biomechanical and histological analyses were performed.

RESULTS

Fourteen eyes were evaluated. There was a statistically significant increase in Young's modulus in Group 1 (102.45%) and in Group 2 (21.30%). In Group 3, no biomechanical changes were measured. Histology showed complete cell loss of keratocytes and endothelium in Group 1 and inhomogeneous keratocyte loss down to 200.0 microm in Group 2. No changes were observed in Group 3.

CONCLUSIONS

Corneal crosslinking without epithelial debridement reduced the biomechanical effect by approximately one fifth compared with standard crosslinking, probably because of restricted and inhomogeneous stromal distribution of riboflavin. The cytotoxic damage was restricted to 200.0 microm stromal depth, which is an advantage over the standard method. Therefore, C3-R is not recommended for the routine treatment of keratoconus but primarily for cases with a corneal thickness less than 400.0 microm in which standard crosslinking cannot be used without serious risk to the endothelium.

OBJECTIVE

Corneal collagen crosslinking (CXL) is designed to halt the progression of keratoconus and corneal ectasia by inducing corneal stiffening. However, it currently is difficult to monitor and evaluate CXL outcome objectively due to the lack of suitable methods to characterize corneal mechanical properties. We validated noncontact Brillouin microscopy to quantify corneal mechanical properties before and after CXL.

METHODS

CXL was performed on fresh porcine eyes using various presoaking times and light doses, with or without epithelial debridement. From Brillouin maps of corneal elastic modulus, stiffness and average modulus of anterior, middle, and posterior stroma were analyzed. Corneal stiffening index (CSI) was introduced as a metric to compare the mechanical efficacy of a given CXL protocol with respect to the standard protocol (30-minute riboflavin presoak, 3 mW/cm² ultraviolet illumination for 30 minutes).

RESULTS

Brillouin corneal stiffness increased significantly (P < 0.001) by epi-off and epi-on CXL. The increase of Brillouin modulus was depth-dependent, indicating that anterior stromal stiffening contributes the most to mechanical outcome. The increase of anterior Brillouin modulus was linearly proportional to the light dose (R²>> 0.98). Compared to the standard epi-off procedure, a typical epi-on procedure resulted in a third of stiffness increase in porcine corneas (CSI = 33).

CONCLUSIONS

Brillouin microscopy allowed imaging and quantifying CXL-induced mechanical changes without contact in a depth-dependent manner at high spatial resolution. This technique may be useful to evaluate the mechanical outcomes of CXL procedures, to compare different crosslinking agents, and for real-time monitoring of CXL in clinical and experimental settings.

Metabolite accumulation has pleiotropic, toxic, or beneficial effects on cell physiology, but such effects are not well understood at the molecular level. Cells respond and adapt to metabolite stress by mechanisms largely unexplored, especially in the context of multiple and simultaneous stresses. Solventogenic and related clostridia have an inherent advantage for production of biofuels and chemicals directly from cellulosic material and other complex carbohydrates, but issues of product/metabolite tolerance and related culture productivities remain. Using DNA microarray-based gene expression analysis, the transcriptional-stress responses of Clostridium acetobutylicum to fermentation acids acetate and butyrate and the solvent product butanol were analyzed and compared in the context of cell physiology. Ontological analysis demonstrated that stress by all three metabolites resulted in upregulation of genes related to post-translational modifications and chaperone activity, and downregulation of the translation-machinery genes. Motility genes were downregulated by acetate-stress only. The general metabolite stress included upregulation of numerous stress genes (dnaK, groES, groEL, hsp90, hsp18, clpC, and htrA), the solventogenic operon aad-ctfA-ctfB, and other solventogenic genes. Acetate stress downregulated expression of the butyryl-CoA- and butyrate-formation genes, while butyrate stress downregulated expression of acetate-formation genes. Pyrimidine-biosynthesis genes were downregulated by most stresses, but purine-biosynthesis genes were upregulated by acetate and butyrate, possibly for thiamine and histidine biosynthesis. Methionine-biosynthesis genes were upregulated by acetate stress, indicating a possibly conserved stress response mechanism also observed in Escherichia coli. Nitrogen-fixation gene expression was upregulated by acetate stress. Butyrate stress upregulated many iron-metabolism genes, riboflavin-biosynthesis genes, and several genes related to cellular repair from oxidative stress, such as perR and superoxide dismutases. Butanol stress upregulated the glycerol metabolism genes glpA and glpF. Surprisingly, metabolite stress had no apparent effect on the expression of the sporulation-cascade genes. It is argued that the list of upregulated genes in response to the three metabolite stresses includes several genes whose overexpression would likely impart tolerance, thus making the information generated in this study, a valuable source for the development of tolerant recombinant strains.

The technique of corneal collagen cross-linking consists of photopolymerization of stromal fibres by the combined action of a photosensitizing substance (riboflavin or vitamin B2) and ultraviolet light from a solid state UVA source. Photopolymerization increases the rigidity of corneal collagen and its resistance to keratectasia. In this report we present two cases, studied through in vivo confocal microscopy, with stage III keratoconus that developed stromal haze after the cross-linking treatment.