Hybrid reconstruction framework for model-based multispectral bioluminescence tomography based on Alpha-divergence

Bioluminescence tomography(BLT)is a promising imaging modality that can provide noninvasive three-dimensional visualization information on tumor distribution.In BLT reconstruction,the widely used methods based on regularization or greedy strategy face problems such as over-sparsity,over-smoothing,spatial discontinuity,poor robustness,and poor multi-target resolution.To deal with these problems,combining the advantages of the greedy strategies as well as regularization methods,we propose a hybrid reconstruction framework for model-based multispectral BLT using the support set of a greedy strategy as a feasible region and the Alpha-divergence to combine the weighted solutions obtained by L1-norm and L2-norm regularization methods.In numerical simulations with digital mouse and in vivo experiments,the results show that the proposed framework has better localization accuracy,spatial resolution,and multi-target resolution.

Study on Shigella Detection by ATP Bioluminescence Magnetic Enzyme Immunoassay

[Objective] The research aimed to establish a rapid detection method for Shigella. [Method] Combining immunomagnetic separation technology with ATP bioluminescence technology, a new kind of fast and accurate ATP bioluminescence magnetic enzyme immunoassay technique for Shigella was established. [Result] Using ATP bioluminescence magnetic enzyme immunoassay technique to detect standard solution for Shigella （ATCC 25931 ）, result showed that correlation coefficient between relative light intensity detected by instrument and bacteria concentration detec- ted by culture counting method was 0.981 1. Moreover, relation curve between relative light intensity and Shigella concentration was drawn. [ Conclusion] The method had a high detection speed and accuracy, and could be used for the rapid detection of pathogen in food and environment.

Rapid bioluminescence assay for monitoring rat CES1 activity and its alteration by traditional Chinese medicines

In traditional Chinese medicine herbs(TCM),including Radix Salviae Miltiorrhizae(Danshen),Radix Puerariae Lobatae(Gegen),Radix Angelicae Sinensis(Danggui),and Rhizoma Chuanxiong(Chuanxiong)are widely used for the prevention and treatment of cardiovascular diseases and also often co-administered with Western drugs as a part of integrative medicine practice.Carboxylesterase 1(CES1)plays a pivotal role in the metabolisms of pro-drugs,Since(S)-2-(2-(6-dimethylamino)-benzothiazole)-4,5-dihydrothiazole-4-carboxylate(NLMe)has recently been identified by us as a selective CES1 bioluminescent sensor,we developed a rapid method using this substrate for the direct measurement of CES1 activity in rats.This bioluminescence assay was applied to determine CES1 activity in rat tissues after a two-week oral administration of each of the four herbs noted above.The results demonstrated the presence of CES1 enzyme in rat blood and all tested tissues with much higher enzyme activity in the blood,liver,kidney and heart than that in the small intestine,spleen,lung,pancreas,brain and stomach.In addition,the four herbs showed tissue-specific effects on rat CES1 expression.Based on the CES1 biodistribution and its changes after treatment in rats,the possibility that Danshen,Gegen and Danggui might alter CES1 activities in human blood and kidney should be considered.In summary,a selective and sensitive bioluminescence assay was developed to rapidly evaluate CES1 activity and the effects of orally administered TCMs in rats.

Source reconstruction for bioluminescence tomography via L_(1/2)regularization

Bioluminescence tomography(BLT)is an important noninvasive optical molecular imaging modality in preclinical research.To improve the image quality,reconstruction algorithms have to deal with the inherent ill-posedness of BLT inverse problem.The sparse characteristic of bioluminescent sources in spatial distribution has been widely explored in BLT and many L1-regularized methods have been investigated due to the sparsity-inducing properties of L1 norm.In this paper,we present a reconstruction method based on L_(1/2) regularization to enhance sparsity of BLT solution and solve the nonconvex L_(1/2) norm problem by converting it to a series of weighted L1 homotopy minimization problems with iteratively updated weights.To assess the performance of the proposed reconstruction algorithm,simulations on a heterogeneous mouse model are designed to compare it with three representative sparse reconstruction algorithms,including the weighted interior-point,L1 homotopy,and the Stagewise Orthogonal Matching Pursuit algorithm.Simulation results show that the proposed method yield stable reconstruction results under different noise levels.Quantitative comparison results demonstrate that the proposed algorithm outperforms the competitor algorithms in location accuracy,multiple-source resolving and image quality.

In vivo bioluminescence imaging of hyperglycemia exacerbating stem cells on choroidal neovascularization in mice

AIM： To investigate the influence of hyperglycemia on the severity of choroidal neovascularization（CNV）,especially the involvement of bone marrow-derived cells（BMCs） and underlying mechanisms.·METHODS： BMCs from firefly luciferase（Fluc）/green fluorescent protein（GFP） double transgenic mice were transplanted into C57BL/6J wide-type mice. The recipient mice were injected intraperitoneally with streptozotocin（STZ） daily for 5 consecutive days to induce diabetes mellitus（DM）, followed by CNV laser photocoagulation.The BMCs recruitment in CNV exposed to hyperglycemia was firstly examined in Fluc/GFP chimeric mice by in vivo optical bioluminescence imaging（BLI） and in vitro Fluc assays. The CNV severity was evaluated by H＆E staining and choroidal flatmount. The expression of vascular endothelial growth factor（VEGF） and stromal cell derived factor-1（SDF-1） was detected by Western blot.·RESULTS： BLI showed that the BMCs exerted dynamic effects in CNV model in Fluc/GFP chimeric mice exposed to hyperglycemia. The signal intensity of transplanted Fluc＋GFP＋BMCs in the DM chimeric mice was significantly higher than that in the control chimeric mice with CNV induction at days 5, 7, 14 and 21（121861.67 ±9948.81 vs 144998.33 ±13787.13 photons/second/cm2/sr for control and DM mice, P5d〈0.05; 178791.67±30350.8 vs240166.67 ±22605.3, P7d〈0.05; 124176.67 ±16253.52 vs196376.67 ±18556.79, P14d〈0.05; 97951.60 ±10343.09 vs119510.00 ±14383.76, P21d〈0.05）, which was consistent with in vitro Fluc assay at day 7 [relative light units of Fluc（RLU1）], 215.00±52.05 vs 707.33±88.65, P 〈0.05; RLU1/relative light units of renilla luciferase（RLU2）, 0.90 ±0.17 vs 1.83 ±0.17, P 〈0.05]. The CNVs in the DM mice were wider than those in the control group at days 5, 7, 14 and21（147.83±17.36 vs 220.33±20.17 μm, P5d〈0.05; 212.17 ±24.63 vs 326.83 ±19.49, P7d〈0.05; 163.17 ±18.24 vs265.17 ±20.55, P14d〈0.05; 132.00 ±10.88 vs 205.33 ±12.98,P21d〈0.05）. The average area of CNV in the DM group was larger at 7d（20688.67±3644.96 vs 32218.00±4132.69 μm2,P 〈0.05）. The expression of VEGF and SDF-1 was enhanced in the DM mice.·CONCLUSION： Hyperglycemia promots the vasculo-genesis of CNV, especially the contribution of BMCs,which might be triggered by VEGF and SDF-1 production.

Three-Dimentional Surface Light Irradiance Reconstruction in Bioluminescence Tomography

Reconstruction of 3D surface irradiance distribution using multiple views captured by charged coupled device(CCD)camera is the basis of solving the light source in bioluminescence tomography(BLT).A simple and convenient mapping technique based on the pin-hole imaging model and Lambert′s cosine law was presented to establish the relationship between gray levels and irradiance intensities.Compared with previous integrating sphere camera calibration used in BLT,the proposed method can effectively avoid heavy burden of simulation experiment to obtain the corresponding relationship of gray levels and irradiance intensities.The accuracy and feasibility of the proposed method are validated with no more than 1mm location error by different types of phantom experiments.The mapping approach is also applicable to other noncontact optical imaging system.

Fast in vivo bioluminescence tomography using a novel pure optical imaging technique

Bioluminescence tomography(BLT)is a novel opt ical molecular imaging technique that advanced the conventional planar bioluminescence imaging(BLI)into a quantifiable three-dimensional(3D)approach in preclinical living animal studies in oncology.In order to solve the inverse problem and reconstruct tumor lesions inside animal body accurately,the prior structural information is com-monly obtained from X ray computed tomography(CT).This strategy requires a complicated hybrid imaging system,extensive post imaging analysis and involvement of ionizing radiation.Moreover,the overall robustness highly depends on the fusion accuracy between the optical and structural information.Here,we present a pure optical bioluminescence tomographic(POBT)system and a novel BLT workfow based on multi-view projection acquisition and 3D surface reconstruction.This met hod can reconstruct the 3D surface of an imaging subject based on a sparse set of planar white-light and bioluminescent images,so that the prior structural information can be offered for 3D tumor lesion reconstruction without the involvement of CT.The performance of this novel technique was evaluated through the comparison with a conventional dual-modality tomo-graphic(DMT)system and a commercialized optical imaging system(IVIS Spectrum)using three breast cancer xenografts.The results revealed that the new technique offered comparable in vivo tomographic accuracy with the DMT system(P>0.05)in much shorter data analysis time.It also offered significantly better accuracy comparing with the IVIS system(P<0.04)without sacrificing too much time.

Influence of the temperature at the Black Sea ctenophores-aliens bioluminescence characteristics

Successful invasion of Mnemiopsis leidyi A. Agassiz, 1865 and Beroe ovata Mayer, 1912 into the Black Sea and their important role in this region pelagic ecosystem is stipulated mainly by the considerable eurythermy of these species. Many ecological-physiological characteristics of ctenophores—aliens are studied quite well. However, bioluminescence, one of the most important elements of the ctenophores ecology and the bioluminescence reaction temperature optimum for these individuals under different environment temperatures were not studied sufficiently. Therefore our researches in this scientific field are significant and conceptually novel for ctenophores ecology study. Experimental investigations were carried out in the period of 2008-2009 in the IBSS. Uni-sized (40 mm) ctenophores were collected in the Sevastopol coastal zone and divided in several groups, contained under different temperatures: from 10°C ± 1°C to 30°C ± 1°C. Ctenophore bioluminescence was investigated under chemical and mechanical stimulation. M. leidyi light emission maximal amplitude (1432.94 ± 71.64 × 108 quantum·s–1·cm–2) with duration of 3.54 ± 0.15 s is fixed under the temperature of 26°C ± 1°C. Temperature increase up to 30°C ± 1°C led to the 4 times decrease of the bioluminescence intensity. Under temperature decrease up to 10°C ± 1°C this parameter decreased 20 times (p B. ovata achieved maximal values under the temperature of 22°C ± 1°C (1150.12 ± 57.51 × 108 quantum·s–1·cm–2) with duration of 3.03 ± 0.15 s. The luminescence intensity decreased under the temperature increase to 30°C ± 1°C more than 20 times. Temperature decrease to the values of 10°C ± 1°C impacted decreasing the amplitude of bioluminescence up to the minimal –4.92 ± 0.22 × 108 quantum·s–1·cm–2. The data obtained testify that characteristics of the ctenophores bioluminescence can be conditioned not only by the modification the environment temperature but by the variability of their physiological condition.

Variability of the bioluminescence characteristics of the Black Sea ctenophores-aliens in connection with different conditions of nutrition

Many ecological-physiological characteristics of ctenophores-aliensMnemiopsis leidyi A. Agassiz, 1865 and Beroe ovata Mayer, 1912 are studied quite well because they play a very important ecological role in the Black Sea ecosystem. However, bioluminescence, one of the most important elements of the ctenophores ecology and its connection with feeding regime were not studied sufficiently. Experiments have shown that characteristics of the ctenophores bioluminescence differed considerably in dependence of food supplies. Thus, amplitude and light-emitting energy of the fed ctenophores B. ovata are maximal, 3 times more than analogical indices of the just-caught individuals and 4 times more than ones of starving individuals. More prolonged flash signal (to 3.5 s), which exceeds light-emitting duration of the starving individuals twice, can be registered from the fed ctenophores. Investigation of the M. leidyi bioluminescence has shown that amplitude and light-emitting energy of the just-caught ctenophores were two times more than those of the starving individuals. At the same time, light-emitting amplitude of the fed individuals is 6.5 times and light-emitting energy is 3 - 4 times higher than that of the just-caught ctenophores. The light-emitting duration of the starving and justcaught organisms is practically the same. The most prolonged signal is registered from the fed ctenophores—up to 2.8 s. The data obtained testify that characteristics of the ctenophores bioluminescence can be conditioned not only by nutritional value but by the composition of the food as well.

Some Theoretical and Applied Aspects of the Oceanic Bioluminescence Registration from Space

The analysis of the surface bioluminescence in the World Ocean registration problems for the purposes of its regions ecological express-monitoring and evaluation of the plankton algae spatial distribution at night time by the existing space systems has been proposed. The connection of the plankton community characteristics with a bioluminescent potential (BP) in the euphotic layer and a possibility to evaluate BP according to bioluminescence intensity in the near-surface layer of 0 - 10 m has been demonstrated. It has been shown that with complete correspondence with the vertical structure in the plankton community at the dark time bioluminescence intensity in 0 - 10 m layer exceeds the same in 60 - 70 m layer for one and a half orders and practically determines BP in 0 - 100 m layer. Peculiarities of the plankton organisms light emission, important for the oceanic bioluminescence registration with the space means of observation are under discussion. Equation for calculation of the measured by the space device sea luminescence level and volume of the bioluminescence intensity in the surface layer of theWorldOcean, which can be registered due to modern technical means of the space systems has been corrected. The conclusion has been done that on the base of the space data about spatial and temporal phytoplankton distribution and bioluminescence “in situ” measurements it is possible to create regional algorithms for transition from numerical estimations of the phytoplankton (chlorophyll “a”) to the day time bioluminescent potential and solution of inverse tasks at night time.

Differential Regulation of Proteins and a Possible Role for Manganese Superoxide Dismutase in Bioluminescence of Panellus stipticus Revealed by Suppression Subtractive Hybridization

Suppression subtractive hybridization (SSH) was employed to investigate bioluminescence in Panellus stipticus (Bull.) P. Karst. by detecting proteins differentially expressed in bioluminescent and luminescent strains. Comparisons of luminescent and non-luminescent monokaryon cultures of North American strains revealed differences in transcript levels of proteins responsible for post-translational modification (PTM) of enzymes. A similar comparison of a luminescent strain of P. stipticus from North America with a non-luminescent European strain revealed the presence of extracellular manganese superoxide dismutase (MnSOD) in the luminescent form, in addition to proteins involved in PTM. The application of MnSOD-specific inhibitors to luminescent mycelium resulted in the rapid loss of luminescence. The relevance to luminescence of proteins involved in PTM is discussed, together with a possible role for MnSOD that considers the potential for SODs to form stable complexes with catechols revealed in previously published research. In light of the recent discovery that hispidine may be the precursor of fungal luciferin, we consider a hypothetical mechanism for fungal luminescence in which the ο-hydroquinone moiety of a hispidine derivative ligates with the extracellular form of MnSOD producing a semiquinone-radical complex, with the resultant semiquinonato complex potentially reacting with molecular oxygen or other reactive oxygen species to produce sufficiently excited intermediates to emit light on relaxation.

Optimizing a high-sensitivity NanoLuc-based bioluminescence system for in vivo evaluation of antimicrobial treatment

Focal and systemic infections are serious threats to human health.Preclinical models enable the development of new drugs and therapeutic regimens.In vivo,animal bioluminescence(BL)imaging has been used with bacterial reporter strains to evaluate antimicrobial treatment effects.However,high-sensitivity bioluminescent systems are required because of the limited tissue penetration and low brightness of the BL signals of existing approaches.Here,we report that NanoLuc(Nluc)showed better performance than LuxCDABE in bacteria.However,the retention rate of plasmid constructs in bacteria was low.To construct stable Staphylococcus aureus reporter strains,a partner protein enolase(Eno)was identified by screening of S.aureus strain USA300 for fusion expression of Nluc-based luciferases,including Nluc,Teluc,and Antares2.Different substrates,such as hydrofurimazine(HFZ),furimazine(FUR),and diphenylterazine(DTZ),were used to optimize a stable reporter strain/substrate pair for BL imaging.S.aureus USA300/Eno-Antares2/HFZ produced the highest number of photons of orange-red light in vitro and enabled sensitive BL tracking of S.aureus in vivo,with sensitivities of approximately 10 CFU from mouse skin and 750 CFU from mouse kidneys.USA300/Eno-Antares2/HFZ was a powerful combination based on the longitudinal evaluation of the therapeutic efficacy of antibiotics.The optimized S.aureus Eno-Antares2/HFZ pair provides a technological advancement for the in vivo evaluation of antimicrobial treatment.

Bio-screening and quantification of methyl paraben in vinegar and coconut juice separated by HPTLC

As a widely used food preservative,methyl paraben was experimentally evidenced with serious hormonelike adverse effects.Herein,a high performance thin-layer chromatography platformed bioluminescent bioautography and image analysis for the selective quantification and confirmation of methyl paraben was proposed and validated in vinegar and coconut juice.First,the detectability of the bioautography to the analyte on different layer materials was estimated,revealing that normal silica gel was the best choice.After that,the liquid of sample extract and working solution were separated to overcome the background noises due to co-extracted matrices.The separation result was then coupled to the optimized bioautography,enabling instant and straightforward screening of the targeted conpound.For accurate quantification,bioluninescent inhibition pattern caused by the analyte was processed by image analysis,giving useful sensitivity(LOD>16 mg/kg),precision(RSD<10.1%)and accuracy(spike-recovery rate 76.9%-112.2%).Finally,the suspected result was confirmed by determining its MS fingerprint,further strengthening the reliability of screening.

Application of stimulating agents on the immobilized bioluminescence strain Pseudomonas putida mt-2 KG1206,preserved by deep-freezing,for the convenient biomonitoring

This study was conducted to develop methods for the application of an immobilized bioluminescence strain （KG1206）, preserved by deep-freezing （DF）, for the monitoring of contaminated environments. The immobilized cells, preserved by DF, required approximately 2 hr for reconstitution of their activity. A large reduction in bioluminescence was observed due to the DF process; 0.07-0.58 times that of the non deep-frozen （NDF） immobilized strain. The decreased bioluminescence activity induced by the DF process was enhanced by the stimulants, sodium lactate （SL） and KNO3. However, regardless of the inducer chemical tested, the immobilized strain modified with KNO3 consistently produced greater bioluminescence than that treated with SL, in the range of 3.0-10.7 （avg. 6.7± 3.69） and 1.2-4.2 （avg. 2.4 ± 1.47） times that of control, respectively. All KNO3 treatments of contaminated groundwater samples also resulted in an increase in bioluminescence activity, but the rate of stimulation varied for each sample. Also, no strong linear correlation was observed between the bioluminescence and the total concentration of an inducer, which may related to the complex characteristics of the environmental samples. Overall, the results demonstrated the ability of immobilized genetically engineered bacteria, preserved by DF, to measure a specific group of environmental contaminants using a stimulating agent （KNO3）, suggesting the potential for its preliminary application in a field-ready bioassay.

Sparse Bayesian reconstruction method for multispectral bioluminescence tomography

We present a sparse Bayesian reconstruction method based on multiple types of a priori information for multispectral bioluminescence tomography （BLT）. In the Bayesian approach, five kinds of a priori information are incorporated, reducing the ill-posedness of BLT. Specifically, source sparsity characteristic is considered to promote reconstruction results. Considering the computational burden in the multispectral case, a series of strategies is adopted to improve computational efficiency, such as optimal permissible source region strategy and node model of the finite element method. The performance of the proposed algorithm is validated by a heterogeneous three-dimensional （3D） micron scale computed tomography atlas and a mouse-shaped phantom. Reconstructed results demonstrate the feasibility and effectiveness of the proposed algorithm.

Improved AFEM algorithm for bioluminescence tomography based on dual-mesh alternation strategy

Adaptive finite element method （AFEM） is broadly adopted to recover the internal source in biological tissues. In this letter, a novel dual-mesh alternation strategy （dual-mesh AFEM） is developed for bioluminescence tomography. By comprehensively considering the error estimation of the finite element method solution on each mesh, two different adaptive strategies based on the error indicator of the reconstructed source and the photon flux density are used alternately in the process. Combined with the constantly adjusted permissible region in the adaptive process, the new algorithm can achieve a more accurate source location compared with the AFEM in the previous experiments.

BIOLUMINESCENCE TOMOGRAPHY:BIOMEDICAL BACKGROUND,MATHEMATICAL THEORY,AND NUMERICAL APPROXIMATION

Over the last couple of years molecular imaging has been rapidly developed to study physiological and pathological processes in vivo at the cellular and molecular levels. Among molecular imaging modalities, optical imaging stands out for its unique advantages, especially performance and cost-effectiveness. Bioluminescence tomography （BLT） is an emerging optical imaging mode with promising biomedical advantages. In this survey paper, we explain the biomedical significance of BLT, summarize theoretical results on the analysis and numerical solution of a diffusion based BLT model, and comment on a few extensions for the study of BLT.

Local Conformational Constraint of Firefly Luciferase Can Affect the Energy of Bioluminescence and Enzyme Stability

Conformational dynamics contribute importantly to enzyme catalysis,such that targeted conformational constraint may affect catalysis.Firefly luciferases undergo extensive structural change during catalysis;key residues form a hydrophobic pocket,excluding water and enabling maximally energetic light production.Point mutants almost always luminesce at longer wavelengths(lower energy)than the wild type.Conformational constraint,using dipeptide analogue 3 at a position critical for optimized excited state structure,produced luciferase emission at a shorter wavelength by∼10 nm.Incomparison,introduction of conformationally constrained analogues 4,5,or 7 afforded luciferases emitting at longer wavelengths,while a related unconstrained luciferase(analogue 6)exhibited wild-type emission.The constrained luciferases tested were more stable than the wild type.Protein modeling demonstrated that the“inside”or“outside”orientation of the conformationally constrained dipeptide led to the shorter or longer emission wavelength,respectively.More broadly,these results suggest that local conformational constraint can control specific elements of enzyme behavior,both in vitro and in vivo.This represents the first example of studying enzyme function by introducing conformationally constrained dipeptides at a specific protein position.The principles discovered here in luciferase modification will enable studies to control the wavelength emission and photophysical properties of modified luciferases.

Joint effects of heavy metal binary mixtures on seed germination,root and shoot growth,bacterial bioluminescence,and gene mutation

This investigation was to assess the joint effects of metal binary mixtures on seed germination,root and shoot growth,bacterial bioluminescence,and gene mutation based on the one toxic unit（1 TU）approach.Different sensitivities and orders of toxicity of metal mixtures were observed among the bioassays.In general,mostly additive or antagonistic effects were observed,while almost no synergistic effects by the binary metal mixtures in all bioassays.Therefore,the combined effects of heavy metals in the different bioassays were difficult to generalize since they were dependent on both chemical type and the organism used in each bioassay.However, these results indicate that a battery of bioassays with mixture chemicals as opposed to just a single assay with single metal is a better strategy for the bioassessment of environmental pollutants.

Moment searching algorithm for bioluminescence tomography

To avoid the ill-posedness in the inverse problem of bioluminescence tomography, a moment searching algorithm fusing the finite element method （FEM） with the moment concept in theoretical mechanics is developed. In the algorithm, the source＇s information is mapped to the surface photon flux density by FEM, and the source＇s position is modified with the feedback through the algorithm of barycenter searching, which makes full use of the position information of the photon flux density on surface. The position is modified in every iterative step and will finally converge to the real source＇s value theoretically.