Optimal Duration Of Exposure Research Articles

Mixed methods study on latent tuberculosis among agate stone workers and advocacy for testing silica dust exposed individuals in India

The 2021 tuberculosis (TB) preventive treatment guidelines in India included silicosis as a screening group, yet latent TB infection (LTBI) testing for silica-dust-exposed individuals is underemphasized. Focusing on an estimated 52 million silica-dust-exposed workers, particularly agate-stone workers in Khambhat, Gujarat, our study aims to estimate LTBI prevalence, identify predictors, and gather insights from TB and silicosis experts. Employing a sequential explanatory mixed-methods approach, a cross-sectional study involved 463 agate-stone workers aged ≥ 20 years in Khambhat, using IGRA kits for LTBI testing. In-depth interviews with experts complemented quantitative findings. Among agate-stone workers, 58% tested positive for LTBI, with predictors including longer exposure, type of work, and BCG vaccination. Our findings reveal a nearly double burden of LTBI compared to the general population, particularly in occupations with higher silica dust exposure. Experts advocate for including silica-dust-exposed individuals in high-risk groups for LTBI testing, exploring cost-effective alternatives like improved skin sensitivity tests, and shorter TB preventive treatment regimens to enhance compliance. Future research should explore upfront TB preventive treatment for silica-dust-exposed individuals with high LTBI prevalence and optimal exposure duration. This study underscores the urgent need for policy changes and innovative approaches to TB prevention among silica-dust-exposed populations, impacting global occupational health strategies.

Biphasic dose-response and effects of near-infrared photobiomodulation on erythrocytes susceptibility to oxidative stress in vitro

The effect of simultaneous application of tert-butyl hydroperoxide (tBHP) and polychromatic near-infrared (NIR) radiation on bovine blood was examined to determine whether NIR light decreases the susceptibility of red blood cells (RBCs) to oxidative stress. The study assessed various exposure methods, wavelength ranges, and optical filtering types. Continuous NIR exposure revealed a biphasic response in cell-free hemoglobin changes, with antioxidative effects observed at low fluences and detrimental effects at higher fluences. Optimal exposure duration was identified between 60 s and 15 min. Protective effects were also tested across wavelengths in the range of 750–1100 nm, with all of them reducing hemolysis, notably at 750 nm, 875 nm, and 900 nm. Comparing broadband NIR and far-red light (750 nm) showed no significant difference in hemolysis reduction. Pulse-dosed NIR irradiation allowed safe increases in radiation dose, effectively limiting hemolysis at higher doses where continuous exposure was harmful. These findings highlight NIR photobiomodulation's potential in protecting RBCs from oxidative stress and will be helpful in the effective design of novel medical therapeutic devices.

NUFIP1-engineered exosomes derived from hUMSCs regulate apoptosis and neurological injury induced by propofol in newborn rats

BackgroundPropofol can increase neurotoxicity in infants but the precise mechanism is still unknown. Our previous study revealed that nuclear FMR1 interacting protein 1 (NUFIP1), a specific ribophagy receptor, can alleviate T cell apoptosis in sepsis. Yet, the effect of NUFIP1-engineered exosomes elicited from human umbilical cord blood mesenchymal stem cells (hUMSCs) on nerve injury induced by propofol remains unclear. This study intended to investigate the effect of NUFIP1-engineered exosomes on propofol-induced nerve damage in neonatal rats. MethodsFirstly, NUFIP1-engineered exosomes were extracted from hUMSCs serum and their identification was conducted using transmission electron microscopy (TEM), Flow NanoAnalyzer, quantitative real-time polymerase chain reaction (qRT-PCR), and western blot (WB). Subsequently, the optimal exposure duration and concentration of propofol induced apoptosis were determined in SH-SY5Y cell line using WB. Following this, we co-cultured the NUFIP1-engineered exosomes in the knockdown group (NUFIP1-KD) and overexpression group (NUFIP1-OE) with SH-SY5Y cells and assessed their effects on the apoptosis of SH-SY5Y cells using terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) assay, Hoechst 33258 staining, WB, and flow cytometry, respectively. Finally, NUFIP1-engineered exosomes were intraperitoneally injected into neonatal rats, and their effects on the learning and memory ability of neonatal rats were observed through the righting reflex and Morris water maze (MWM) test. Hippocampi were extracted from different groups for hematoxylin-eosin (HE) staining, immunohistochemistry, immunofluorescence, and WB to observe their effects on apoptosis in neonatal rats. ResultsTEM, Flow NanoAnalyzer, qRT-PCR, and WB analyses confirmed that the exosomes extracted from hUMSCs serum exhibited the expected morphology, diameter, surface markers, and expression of target genes. This confirmed the successful construction of NUFIP1-KD and NUFIP1-OE-engineered exosomes. Optimal exposure duration and concentration of propofol were determined to be 24 hours and 100 µg/ml, respectively. Co-culture of NUFIP1 engineered exosomes and SH-SY5Y cells resulted in significant up-regulation of pro-apoptotic proteins Bax and c-Caspase-3 in the KD group, while anti-apoptotic protein Bcl-2 was significantly decreased. The OE group showed the opposite trend. TUNEL apoptosis assay, Hoechst 33258 staining, and flow cytometry yielded consistent results. Animal experiments demonstrated that intraperitoneal injection of NUFIP1-KD engineered exosomes prolonged the righting reflex recovery time of newborn rats, and MWM tests revealed a significant diminution in the time and number of newborn rats entering the platform. HE staining, immunohistochemistry, immunofluorescence, and WB results also indicated a significant enhancement in apoptosis in this group. Conversely, the experimental results of neonatal rats in the OE group revealed a certain degree of anti-apoptotic effect. ConclusionsNUFIP1-engineered exosomes from hUMSCs have the potential to regulate nerve cell apoptosis and mitigate neurological injury induced by propofol in neonatal rats. Targeting NUFIP1 may hold great significance in ameliorating propofol-induced nerve injury.

The potential of Euglena species as a bioindicator for soil ecotoxicity assessment

Currently, there are no standard international test methods for assessing aquatic and soil toxicity, with aquatic toxicity tests based on limited Euglena species. Here, we proposed Euglena species as extended test species, especially as new soil test species for a paper-disc soil method, considering its ecologically important roles in providing highly bioavailable in-vivo nutrients to upper trophic level organisms. We conducted experiments to identify the optimal exposure duration for two Euglena species (Euglena viridis and Euglena geniculata). We demonstrated the toxic effects of nickel (model contaminant) on their photosynthetic parameters and growth in freshwater. The growth and photosynthetic activity of three Euglena species were significantly inhibited in nickel-contaminated soil during paper-disc soil tests, especially the test species adsorbed onto paper-disc soil. Euglena gracilis was more sensitive to nickel than E. viridis and E. geniculata in freshwater and soil. Thus, E. viridis and E. geniculata have potential as additional test species for improving test species diversity, while all three species have potential as new soil test species for soil toxicity assessment. Thus, results these species may be suitable for routine aquatic toxicity testing and new soil toxicity testing, addressing the current paucity of test species in freshwater and soil toxicity assessment.

Effectiveness of endometrial scratching in infertility treatment with assisted reproductive technologies

Embryo implantation is a key factor limiting the effectiveness of assisted reproductive technologies, and depends on the quality of the blastocyst and the susceptibility of the endometrium. Various methods of treatment aimed at improving the receptivity of the uterine mucosa, including endometrial scratching, have been proposed. At the same time, the main mechanisms underlying this intervention remain unclear. Moreover, the optimal duration of exposure to the uterine mucosa and the groups of patients in which it can have a positive effect have not been determined. The study aims to investigate the main mechanisms and evaluate the effectiveness of local endometrial damage performed at various times before embryo transfer in the program of assisted reproductive technologies in women with and without a history of implantation failures based on the analysis of scientific literature. Clinical studies, systematic reviews and meta-analyses were studied in PubMed, Embase, Cochrane Library, Web of Science databases, Elsevier, Springer, Wiley publishers, on eLibrary, Research Gate platforms, in open sources of the scientific and educational segment of the Internet. The literature search was conducted by keywords: endometrial scratching, local endometrial damage, implantation failures, endometrial receptivity, assisted reproductive technologies, infertility. The search depth is 20 years. The main mechanisms of the effect of scratching on increasing endometrial receptivity include decidualization, the development of a local inflammatory reaction, as well as improved synchronization between the uterine mucosa and the transferred embryo by preventing premature maturation of the endometrium. Studies that showed a positive effect of local damage to the uterine mucosa were mainly conducted in patients with repeated implantation failures and with intervention in the luteal phase of the cycle preceding stimulation. At the same time, the work performed in a non-selective population of women and non-standardized by exposure time, in most cases, did not reveal the advantages of endometrial scratching. In this regard, additional qualitative randomized controlled trials with a large sample size and a clear identification of groups of patients for whom this intervention may benefit are required.

Prolonging the time of progesterone supplementation to improve the pregnancy outcomes of single day 6 blastocyst transfer in frozen-thawed cycles: study protocol for a randomized controlled trial

BackgroundInfertility is one of the most important and underappreciated reproductive health problems in developing countries. Currently, in vitro fertilization and embryo transfer is the most effective treatment strategy for infertility. In a frozen-thawed cycle, single-blastocyst transfer can not only ensure relatively higher pregnancy and live birth rates but also effectively reduce the risk of maternal and neonatal complications. In frozen-thawed cycles, progesterone is initiated to promote the final phase of endometrial preparation prior to embryo transfer. However, the optimal duration of exposure to progesterone has remained inconclusive. Therefore, we designed a randomized controlled trial (RCT) to compare the effects of different prolonged progesterone transformation times (P+6 and P+7) on the pregnancy outcomes of D6 single blastocyst transfer in a frozen-thawed cycle.MethodsThis is a single-center, prospective, randomized controlled clinical trial involving 900 patients with single blastocyst transfer in the frozen-thawed cycle, aged from 20 to 38 years, with less than three transfers, and with HRT-cycle single D6 blastocyst transfer in the current cycle. Participants will be randomly assigned (1:1) into two parallel groups: the transfer of day 6 blastocysts on the 7th day of progesterone supplementation and the transfer of day 6 blastocysts on the 6th day of progesterone supplementation. The primary outcome measure is the clinical pregnancy rate. Secondary outcome measures include the miscarriage rate and live birth rate.DiscussionThis is the first randomized controlled trial to compare the transfer of day 6 blastocysts on the 6th and 7th day of progesterone supplementation. The results of this study will provide evidence for whether to prolong the duration of exposure to progesterone prior to embryo transfer.Trial registrationClinicalTrials.gov, ID: NCT04938011. Registered on 19 June 2021.

Transport Characteristics of Plasma Membranes of PK-15 Passaged Cells

The values of permeability coefficients to water molecules and cryoprotectants are demanded to select the optimal duration of exposure of cells in cryoprotective media at the stage of their preparation for cryopreservation, as well as to find optimal cooling and warming rates during the freeze-thawing of cell suspensions. The necessary numerical values of such cell parameters as the osmotically inactive volume α and the surface-area-to-volume ratio γ were obtained for the analytical evaluation of the permeability coefficients of the PK-15 cells’ plasma membranes using physico-mathematical modelling. The permeability coefficients kp of the plasma membranes of PK-15 cells to 1,2-PD, EG, DMSO and glycerol cryoprotectants molecules, as well as the filtration coefficients Lp to water molecules at temperatures of 25, 15 and 5°C were determined by approximating the experimental data of the change in relative volume of cells on exposure time in the studied solutions by theoretical curves calculated on the basis of physical and mathematical model of passive transport of water and permeable substances under the condition of their maximum coincidence. The value of the activation energy of the transmembrane transfer of molecules of these substances is calculated

How varying parameters impact insecticide resistance bioassay: An example on the worldwide invasive pest Drosophila suzukii.

Monitoring pesticide resistance is essential for effective and sustainable agricultural practices. Bioassays are the basis for pesticide-resistance testing, but devising a reliable and reproducible method can be challenging because these tests are carried out on living organisms. Here, we investigated five critical parameters and how they affected the evaluation of resistance to the organophosphate phosmet or the pyrethroid lambda-cyhalothrin using a tarsal-contact protocol on Drosophila suzukii, a worldwide invasive pest. Three of the parameters were related to insect biology: (i) sex, (ii) age of the imago (adult stage) and (iii) genetic diversity of the tested population. The two remaining parameters were linked to the experimental setup: (iv) the number of individuals tested per dose and (v) the duration of exposure to the active ingredient. Results showed that response to insecticide differed depending on sex, males being twice as susceptible to phosmet as females. Age principally affected young females' susceptibility to phosmet, because 0-24 hour-old flies were twice as susceptible as 24-48 hour-old and 72-96 hour-old females. Genetic diversity had no observable effect on resistance levels. The precision and accuracy of the median lethal dose (LD50) were greatly affected by the number of individuals tested per dose with a threshold effect. Finally, optimal duration of exposure to the active ingredient was 24 h, as we found an underestimation of mortality when assessed between 1 and 5 h after exposure to lambda-cyhalothrin. None of the main known point mutations on the para sodium channel gene associated with a knockdown effect were observed. Our study demonstrates the importance of calibrating the various parameters of a bioassay to develop a reliable method. It also provides a valuable and transferable protocol for monitoring D. suzukii resistance worldwide.

The effects of Virtual Reality technology application in the aircraft pilot training process

In this study, we focus on some problems related to virtual reality technology usage in supporting the educational process. The main goal is to obtain the optimal duration of exposure to virtual reality without the appearance of physiological problems for the user affected the achievement of teaching outcomes. Most of the characteristics of virtual reality software at the same time point out some issues in the areas of technological innovations, especially in the health context. The presented example of the VR environment’s training application shows the effectiveness of VR methods and their possibilities in training aircraft pilots. Teaching involving VR allowed to achieve, on average, as much as 90% correct answers, which in comparison with traditional methods resulted in an increase of 21 to 32.7% of correct answers in comparison with training using VR. The research confirmed that a critical challenge in the context of the broader use of VR in education is to appropriately adapt the learning materials and content to the technical requirements. Then, in certain areas, the use of VR is beneficial.

Bidirectional myofiber transition through altering the photobiomodulation condition

Despite remarkable advancements in modern medicine, muscular atrophy remains as an unsolved problem. It is well known that pathological characteristics of different atrophy types could vary according to the pathophysiological causes. In fact, the lesion of atrophy is not always homogenously distributed but often predominantly evident in either fast or slow myofibers. As the focalization of the atrophic lesions, the existence and the functional impairment of each fast and slow progenitor/satellite cell (SC) are suspected though there are still controversies about this hypothesis. In this study, we isolated Pax7 positive (Pax7+ve) SCs from the tibia anterior (fast) and soleus (slow) muscles respectively and successfully demonstrated, for the first time, the difference between optimal exposure durations of photobiomodulation (PBM) which was known as low level laser irradiation (LLLI) in promoting proliferation of Pax7+ve SC which were acquired from fast and slow muscles respectively. Moreover, a hypertrophy-accompanied bidirectional change in myofiber composition with neuromuscular junction alteration, either from slow to fast or fast to slow, were achieved by applying different PBM durations. Simultaneously, PBM exhibited a synergistic effect with muscle exercise on the increase in myofiber size. Our data suggested the existence of at least two different populations of Pax7+ve SC which possess distinct sensitivities towards PBM. As our data revealed the capability of PBM in bidirectional changes of skeletal muscle composition and neuromuscular junction constitution thereby strengthen its contractility through altering the irradiation condition, we believe PBM showed the potential to be as a promising clinical treatment for muscular atrophy.

The relation of Bleomycin Delivery Efficiency to Microbubble Sonodestruction and Cavitation Spectral Characteristics

The concurrent assessment of principal sonoporation factors has been accomplished in a single systemic study. Microbubble sonodestruction dynamics and cavitation spectral characteristics, ultrasound scattering and attenuation, were examined in relation to the intracellular delivery of anticancer drug, bleomycin. Experiments were conducted on Chinese hamster ovary cells coadministered with Sonovue microbubbles. Detailed analysis of the scattering and attenuation temporal functions culminated in quantification of metrics, inertial cavitation dose and attenuation rate, suitable for cavitation control. The exponents, representing microbubble sonodestruction kinetics were exploited to derive dosimetric, microbubble sonodestruction rate. High intracorrelation between empirically-attained metrics defines the relations which indicate deep physical interdependencies within inherent phenomena. Subsequently each quantified metric was validated to be well-applicable to prognosticate the efficacy of bleomycin delivery and cell viability, as indicated by strong overall correlation (R2 > 0.85). Presented results draw valuable insights in sonoporation dosimetry and contribute towards the development of universal sonoporation dosimetry model. Both bleomycin delivery and cell viability reach their respective plateau levels by the time, required to attain total microbubble sonodestruction, which accord with scattering and attenuation decrease to background levels. This suggests a well-defined criterion, feasible through signal-registration, universally employable to set optimal duration of exposure for efficient sonoporation outcome.

Assessment of metal/metalloid occurrence in rivers with their accumulation in macrophyte case study with Myriophyllum alterniflorum.

Water quality monitoring with integrative tools is a main issue of concern for environment assessment. Submerged aquatic macrophyte can be a good candidate for the evaluation of contaminant content in rivers. Indeed, owing to their habitat, aquatic macrophytes interact directly with surface water; they can absorb contaminants and thus allow to detect their presence in water. In situ studies were conducted over 28days in five aquatic environments, affected by different levels of anthropogenic pressure (domestic wastewater plant, industrial activities), during two field campaigns. We have investigated whether the accumulation of some metals and a metalloid (As) in Myriophyllum alterniflorum could be used to detect their occurrence in river waters. Our results demonstrated that long time bioaccumulation was correlated with the contaminant levels in water. However, the water composition and the duration of exposure affected the studied pollutants' absorption. On an operational point of view, the optimal duration of exposure of Myriophyllum alterniflorum to assess the water quality is conditioned by the contaminant contents in waters that can induce different defense mechanisms as the reduction of pollutant absorption and their efflux. In addition, the nutrient concentration influenced the accumulation of pollutants since the higher the nutrient level, the higher the essential metal accumulation was observed.

Exposure Duration Is a Determinant of the Effect of Sinusoidal Electromagnetic Fields on Peak Bone Mass of Young Rats.

We proposed a three-step strategy to obtain the optimal therapeutic parameters, which is composed of large-scale screening at cellular level, verification in animal experiments, and confirmation by a clinical trial. The objective of the current study was to test the feasibility of our strategy. Newborn rat calvarial osteoblasts were treated by 50Hz 1.8mT sinusoidal electromagnetic fields (SEMFs) with 0.5, 1.0, 1.5, 2.0, 2.5, and 3.0h/days, respectively. The osteogenic differentiation and maturation of the osteoblast were assayed and compared to obtain the optimal duration. One-month-old growing rats were then treated by the same SEMFs with 0.5, 1.5, and 2.5h/days, respectively, and the peak bone mass was analyzed after 2months. It was found that the optimal exposure duration to promote the osteogenic differentiation and maturation of osteoblasts was 1.5h/days, judging by the increasing degrees of ALP activity, calcified nodules formed, the gene and protein expression levels of Runx-2, BMP-2, and Col-I, as well as the expression levels of signaling proteins of the BMP-2/Smad1/5/8 pathway. The highest increase of peak bone mass after 2months was also obtained by 1.5h/days, judging by the results of X-ray dual-energy absorptiometry, mechanical property analysis, micro-CT scanning, and serum bone turnover marker examinations. The above results indicated that exposure duration is a determinant for the therapeutic effect of EMFs, and the optimal therapeutic effects only can be obtained by the optimal exposure duration.

Acoustic Control of Mosquito Larvae In Artificial Drinking Water Containers

Emerging technology designed to kill mosquito larvae with sound waves may present a nonchemical and nonbiological alternative to reduce larval populations of key medically important mosquito species such as Aedes aegypti in containers or catchments of water. These devices could benefit integrated vector management programs facing public resistance to the use of chemical or biological larvicides in stored drinking water. In this study we investigate the efficacy of a Larvasonic SD-Mini Acoustic Larvicide device in reducing larval populations of Ae. aegypti in 3 volumes of water across a range of acoustic exposure durations. We report lethal pulse duration times for 50% and 90% mortality and optimal exposure durations for the tested water volumes.

Investigation of Microbubble Cavitation-Induced Calcein Release from Cells In Vitro

In the present study, microbubble (MB) cavitation signal analysis was performed together with calcein release evaluation in both pressure and exposure duration domains of the acoustic field. A passive cavitation detection system was used to simultaneously measure MB scattering and attenuation signals for subsequent extraction efficiency relative to MB cavitation activity. The results indicate that the decrease in the efficiency of extraction of calcein molecules from Chinese hamster ovary cells, as well as cell viability, is associated with MB cavitation activity and can be accurately predicted using inertial cavitation doses up to 0.18 V × s (R2 > 0.9, p < 0.0001). No decrease in additional calcein release or cell viability was observed after complete MB sonodestruction was achieved. This indicates that the optimal exposure duration within which maximal sono-extraction efficiency is obtained coincides with the time necessary to achieve complete MB destruction. These results illustrate the importance of MB inertial cavitation in the sono-extraction process. To our knowledge, this study is the first to (i) investigate small molecule extraction from cells via sonoporation and (ii) relate the extraction process to the quantitative characteristics of MB cavitation acoustic spectra.

Optimal preparation of the complex of paclitaxel loaded heparin nanoparticles and microbubbles and screening for the optimized ultrasound condition for different breast cancer cells

Objective To investigate the optimal preparation of the complex of paclitaxel-loaded heparin nanoparticles and microbubbles and screen the optimized ultrasound (US) condition for highly aggressive breast cancer cells MDA-MB-231 and less aggressive breast cancer cells MCF-7. Methods Paclitaxel-loaded nanoparticles were prepared using heparin as nanocarrier, and biotinlyated microbubbles were prepared by mechanical shaking. Paclitaxel-loaded nanoparticles was conjugated with microbubbles to prepare the complex using avidin-biotin system. The optimized method was screened. Breast cancer cell MDA-MB-231 and MCF-7 were treated with the prepared complex and different ultrasonic parameters including irradiation intensity and exposure duration were evaluated by cellular uptake experiments. Results The drug loading in the complex was 106.7 μg/108 microbubbles when the paclitaxel-nanoparticle concentration was 2 mg/ml. For MDA-MB-231 cells, the optimal irradiation intensity was 1.0 W/cm2 and the optimal exposure duration was 40 s, and they were 1.5W/cm2 and 40s for MCF-7 cells, respectively. Conclusions The complex with high drug loading weight, composed of paclitaxel-loaded heparin nanoparticles and microbubbles, was successfully prepared, and it was found the optimal ultrasound condition was different in two breast cancer cells. Key words: Ultrasonography; Paclitaxel-nanoparticle; Microbubbles; Tumor cells

Optimization of Chemical Inhibition Conditions for NMD Studies in p53‐Positive Cancer Cell Lines (LB234)

We report on the treatment of p53‐positive cancer cell lines with chemical inhibitors of transcription, translation and PI‐3 kinase‐like kinases (PIKKs). Our goal was to determine optimal concentrations of, and exposure times for, chemical inhibitor treatments. These experiments were prerequisite for our studies of the role nonsense‐mediated mRNA decay (NMD) plays in p53 expression. Cultured p53‐positive cancer cells were treated with varied concentrations of cycloheximide, caffeine, wortmannin, or actinomycin D to determine optimal concentrations. Cells were also treated with constant inhibitor concentrations in time‐course experiments to determine optimal duration of exposure. Treated cell samples were divided equally for protein and total RNA isolation. Protein samples were subjected to SDS‐PAGE and western analyses for p53 and beta‐actin. RNA samples were used to synthesize cDNA, which was subjected to TaqMan quantitative RT‐PCR (qRT‐PCR) analysis for p53 and beta‐actin. Western and qRT‐PCR data are consistent with optimal chemical inhibitor concentrations being substantially higher for HepG2 cells compared to MCF‐7 and HeLa. However, chemical inhibitor exposure times required to achieve optimal effect was not widely varied between cell lines studied. Through our determination of optimal conditions for chemical inhibition of transcription, translation and PIKK activity in p53‐positive cell lines, we conclude that cell lines studied require widely different concentrations of, but not different exposure times to, chemical inhibitors. Therefore, we determined optimal concentrations and exposure times for chemical inhibition in our studies of the role of NMD in p53 expression.Grant Funding Source: University of Tampa Chemistry, Biochemistry &amp; Physics Departmental Student Research Support Fund

Histological and Biochemical Analysis of Mechanical and Thermal Bioeffects in Boiling Histotripsy Lesions Induced by High Intensity Focused Ultrasound

Recent studies have shown that shockwave heating and millisecond boiling in high-intensity focused ultrasound fields can result in mechanical fractionation or emulsification of tissue, termed boiling histotripsy. Visual observations of the change in color and contents indicated that the degree of thermal damage in the emulsified lesions can be controlled by varying the parameters of the exposure. The goal of this work was to examine thermal and mechanical effects in boiling histotripsy lesions using histologic and biochemical analysis. The lesions were induced in ex vivo bovine heart and liver using a 2-MHz single-element transducer operating at duty factors of 0.005–0.01, pulse durations of 5–500 ms and in situ shock amplitude of 73 MPa. Mechanical and thermal damage to tissue was evaluated histologically using conventional staining techniques (hematoxylin and eosin, and nicotinamide adenine dinucleotide-diaphorase). Thermal effects were quantified by measuring denaturation of salt soluble proteins in the treated region. According to histologic analysis, the lesions that visually appeared as a liquid contained no cellular structures larger than a cell nucleus and had a sharp border of one to two cells. Both histologic and protein analysis showed that lesions obtained with short pulses (<10 ms) did not contain any thermal damage. Increasing the pulse duration resulted in an increase in thermal damage. However, both protein analysis and nicotinamide adenine dinucleotide-diaphorase staining showed less denaturation than visually observed as whitening of tissue. The number of high-intensity focused ultrasound pulses delivered per exposure did not change the lesion shape or the degree of thermal denaturation, whereas the size of the lesion showed a saturating behavior suggesting optimal exposure duration. This study confirmed that boiling histotripsy offers an effective, predictable way to non-invasively fractionate tissue into sub-cellular fragments with or without inducing thermal damage.

Effect of seaweed extracts and plant growth regulators on high-frequency in vitro mass propagation of Lycopersicon esculentum L (tomato) through double cotyledonary nodal explant

An efficient and reproducible two-step in vitro propagation system for tomato (Lycopersicon esculentum) was developed by using the combinations of seaweed biostimulant (Gracilaria edulis and Sargassum wightii) extracts and plant growth regulators (PGRs). Double cotyledonary nodal (DCN) explants of Co-3 cultivar were initially cultured on Murashige and Skoog (MS) and Gamborg’s medium (B5) containing thidiazuron (TDZ) and 6-benzylaminopurine (BA); the best responding cytokinin was tested in combinations with different auxins (NAA, IAA and IBA), and seaweed extracts (G. edulis and S. wightii) of about basal MS medium +10–70% was used for shoot proliferation. The best organogenic culture response was obtained on MS medium fortified with 1.5 mg L−1 TDZ and 1.5 mg L−1 IBA. Up to 24 shoots per explants were formed at an optimal duration of exposure to 35 days. Mini shoots of about 3–4 cm were transferred to medium supplemented with MS + iP, MS + zeatin, MS + G. edulis and MS + S. wightii at different concentrations. High frequency of shoot elongation was observed in the medium supplemented with 30% G. edulis (15.2 cm), and profuse rooting was observed in the medium supplemented with 50% S. wightii of about 16.1 cm. Shoot elongation and rooting were observed in the medium supplemented with seaweed extracts. The plantlets were transferred to the plant growth chamber (70% of relative humidity and 9 light cycles) and maintained in it for a week, and then they were transferred to a greenhouse condition. The plant growth chamber to green house transferred plantlets showed an increase in the survival rate from 70 to 85%. Thus a two-step regeneration protocol was developed in this study with a combination of seaweed extracts and PGRs, which provides a basis for the production of transgenics with high frequency and survivability of tomato plants.

A Combined Experimental and Mathematical Approach for Molecular-based Optimization of Irinotecan Circadian Delivery

Circadian timing largely modifies efficacy and toxicity of many anticancer drugs. Recent findings suggest that optimal circadian delivery patterns depend on the patient genetic background. We present here a combined experimental and mathematical approach for the design of chronomodulated administration schedules tailored to the patient molecular profile. As a proof of concept we optimized exposure of Caco-2 colon cancer cells to irinotecan (CPT11), a cytotoxic drug approved for the treatment of colorectal cancer. CPT11 was bioactivated into SN38 and its efflux was mediated by ATP-Binding-Cassette (ABC) transporters in Caco-2 cells. After cell synchronization with a serum shock defining Circadian Time (CT) 0, circadian rhythms with a period of 26 h 50 (SD 63 min) were observed in the mRNA expression of clock genes REV-ERBα, PER2, BMAL1, the drug target topoisomerase 1 (TOP1), the activation enzyme carboxylesterase 2 (CES2), the deactivation enzyme UDP-glucuronosyltransferase 1, polypeptide A1 (UGT1A1), and efflux transporters ABCB1, ABCC1, ABCC2 and ABCG2. DNA-bound TOP1 protein amount in presence of CPT11, a marker of the drug PD, also displayed circadian variations. A mathematical model of CPT11 molecular pharmacokinetics-pharmacodynamics (PK-PD) was designed and fitted to experimental data. It predicted that CPT11 bioactivation was the main determinant of CPT11 PD circadian rhythm. We then adopted the therapeutics strategy of maximizing efficacy in non-synchronized cells, considered as cancer cells, under a constraint of maximum toxicity in synchronized cells, representing healthy ones. We considered exposure schemes in the form of an initial concentration of CPT11 given at a particular CT, over a duration ranging from 1 to 27 h. For any dose of CPT11, optimal exposure durations varied from 3h40 to 7h10. Optimal schemes started between CT2h10 and CT2h30, a time interval corresponding to 1h30 to 1h50 before the nadir of CPT11 bioactivation rhythm in healthy cells.