The optimal measurement period of actigraphy for circadian rhythm in relation to adiposity: A retrospective case-control study.

BACKGROUND: This study focused on the relationship between adiposity and Rest-Activity Rhythms (RAR), utilizing both parametric cosine-based models and non-parametric algorithms. The emphasis was on the impact of varying measurement periods (7-28 days) on this relationship. METHODS: We retrieved actigraphy data from two datasets, encompassing a diverse cohort recruited from an obesity outpatient clinic and a workplace health promotion program. Participants were required to wear a research-grade wrist actigraphy device continuously for a minimum of four weeks. The final dataset included 115 individuals (mean age 40.7 ± 9.5 years, 51 % female). We employed both parametric and non-parametric methods to quantify RAR using six standard variables. Additionally, the study evaluated the correlations between three key adiposity indices - Body Mass Index (BMI), Visceral Adipose Tissue (VAT) area, and Body Fat Percentage (BF%) - and circadian rhythm indicators, controlling for factors like physical activity, age, and gender. RESULTS: The obesity group displayed a significantly lower relative amplitude (RA) as per non-parametric algorithm findings, with a decreased amplitude noted in the parametric algorithm analysis, in comparison to the overweight and control groups. The relationship between circadian rhythm indicators and adiposity metrics over 7- to 28-day periods was examined. A notable negative correlation was observed between RA and both BMI and VAT, while correlation coefficients between adiposity indicators and non-parametric circadian parameters increased with extended durations of actigraphy data. Specifically, RA over a 28-day period was significantly correlated with BF%, a trend not seen in the 7-day measurement (p = 0.094) in multivariate linear regression. The strength of the correlation between BF% and 28-day RA was more pronounced than that in the 7-day period (p = 0.044). However, replacing RA with amplitude as per parametric cosinor fitting yielded no significant correlations for any of the measurement periods. CONCLUSION: The study concludes that a 28-day measurement period more effectively captures the link between disrupted circadian rhythms and adiposity. Non-parametric algorithms, in particular, were more effective in characterizing disrupted circadian rhythms, especially when extending the measurement period beyond the standard 7 days.

Increase of HCN current in SOD1-associated amyotrophic lateral sclerosis.

The clinical manifestations of sporadic amyotrophic lateral sclerosis (ALS) vary widely. However, the current classification of ALS is mainly based on clinical presentations, while the roles of electrophysiological and biomedical biomarkers remain limited. Herein, we investigated a group of patients with sporadic ALS and an ALS mouse model with superoxide dismutase 1 (SOD1)/G93A transgenes using nerve excitability tests (NET) to investigate axonal membrane properties and chemical precipitation, followed by enzyme-linked immunosorbent assay analysis to measure plasma misfolded protein levels. Six of 19 patients (31.6%) with sporadic ALS had elevated plasma misfolded SOD1 protein levels. In sporadic ALS patients, only those with elevated misfolded SOD1 protein levels showed an increased inward rectification in the current-threshold (I/V) curve and an increased threshold reduction in the hyperpolarizing threshold electrotonus (TE) in the NET study. Two familial ALS patients with SOD1 mutations also exhibited similar electrophysiological patterns of NET. For patients with sporadic ALS showing significantly increased inward rectification in the I/V curve, we noted an elevation in plasma misfolded SOD1 level, but not in total SOD1, misfolded C9orf72, or misfolded phosphorylated TDP43 levels. Computer simulations demonstrated that the aforementioned axonal excitability changes are likely associated with an increase in hyperpolarization-activated cyclic nucleotide-gated (HCN) current. In SOD1/G93A mice, NET also showed an increased inward rectification in the I/V curve, which could be reversed by a single injection of the HCN channel blocker, ZD7288. Daily treatment of SOD1/G93A mice with ZD7288 partially prevented the early motor function decline and spinal motor neuron death. In summary, sporadic ALS patients with elevated plasma misfolded SOD1 exhibited similar patterns of motor axonal excitability changes as familial ALS patients and ALS mice with mutant SOD1 genes, suggesting the existence of SOD1-associated sporadic ALS. The observed NET pattern of increased inward rectification in the I/V curve was attributable to an elevation in the HCN current in SOD1-associated ALS.

Rest-Activity Rhythm Differences in Acute Rehabilitation Between Poststroke Patients and Non-Brain Disease Controls: Comparative Study.

BACKGROUND: Circadian rhythm disruptions are a common concern for poststroke patients undergoing rehabilitation and might negatively impact their functional outcomes. OBJECTIVE: Our research aimed to uncover unique patterns and disruptions specific to poststroke rehabilitation patients and identify potential differences in specific rest-activity rhythm indicators when compared to inpatient controls with non-brain-related lesions, such as patients with spinal cord injuries. METHODS: We obtained a 7-day recording with a wearable actigraphy device from 25 poststroke patients (n=9, 36% women; median age 56, IQR 46-71) and 25 age- and gender-matched inpatient control participants (n=15, 60% women; median age 57, IQR 46.5-68.5). To assess circadian rhythm, we used a nonparametric method to calculate key rest-activity rhythm indicators-relative amplitude, interdaily stability, and intradaily variability. Relative amplitude, quantifying rest-activity rhythm amplitude while considering daily variations and unbalanced amplitudes, was calculated as the ratio of the difference between the most active 10 continuous hours and the least active 5 continuous hours to the sum of these 10 and 5 continuous hours. We also examined the clinical correlations between rest-activity rhythm indicators and delirium screening tools, such as the 4 A's Test and the Barthel Index, which assess delirium and activities of daily living. RESULTS: Patients who had a stroke had higher least active 5-hour values compared to the control group (median 4.29, IQR 2.88-6.49 vs median 1.84, IQR 0.67-4.34; P=.008). The most active 10-hour values showed no significant differences between the groups (stroke group: median 38.92, IQR 14.60-40.87; control group: median 31.18, IQR 18.02-46.84; P=.93). The stroke group presented a lower relative amplitude compared to the control group (median 0.74, IQR 0.57-0.85 vs median 0.88, IQR 0.71-0.96; P=.009). Further analysis revealed no significant differences in other rest-activity rhythm metrics between the two groups. Among the patients who had a stroke, a negative correlation was observed between the 4 A's Test scores and relative amplitude (ρ=-0.41; P=.045). Across all participants, positive correlations emerged between the Barthel Index scores and both interdaily stability (ρ=0.34; P=.02) and the most active 10-hour value (ρ=0.42; P=.002). CONCLUSIONS: This study highlights the relevance of circadian rhythm disruptions in poststroke rehabilitation and provides insights into potential diagnostic and prognostic implications for rest-activity rhythm indicators as digital biomarkers.

Comparing Human-Smartphone Interactions and Actigraphy Measurements for Circadian Rhythm Stability and Adiposity: Algorithm Development and Validation Study.

BACKGROUND: This study aimed to investigate the relationships between adiposity and circadian rhythm and compare the measurement of circadian rhythm using both actigraphy and a smartphone app that tracks human-smartphone interactions. OBJECTIVE: We hypothesized that the app-based measurement may provide more comprehensive information, including light-sensitive melatonin secretion and social rhythm, and have stronger correlations with adiposity indicators. METHODS: We enrolled a total of 78 participants (mean age 41.5, SD 9.9 years; 46/78, 59% women) from both an obesity outpatient clinic and a workplace health promotion program. All participants (n=29 with obesity, n=16 overweight, and n=33 controls) were required to wear a wrist actigraphy device and install the Rhythm app for a minimum of 4 weeks, contributing to a total of 2182 person-days of data collection. The Rhythm app estimates sleep and circadian rhythm indicators by tracking human-smartphone interactions, which correspond to actigraphy. We examined the correlations between adiposity indices and sleep and circadian rhythm indicators, including sleep time, chronotype, and regularity of circadian rhythm, while controlling for physical activity level, age, and gender. RESULTS: Sleep onset and wake time measurements did not differ significantly between the app and actigraphy; however, wake after sleep onset was longer (13.5, SD 19.5 minutes) with the app, resulting in a longer actigraphy-measured total sleep time (TST) of 20.2 (SD 66.7) minutes. The obesity group had a significantly longer TST with both methods. App-measured circadian rhythm indicators were significantly lower than their actigraphy-measured counterparts. The obesity group had significantly lower interdaily stability (IS) than the control group with both methods. The multivariable-adjusted model revealed a negative correlation between BMI and app-measured IS (P=.007). Body fat percentage (BF%) and visceral adipose tissue area (VAT) showed significant correlations with both app-measured IS and actigraphy-measured IS. The app-measured midpoint of sleep showed a positive correlation with both BF% and VAT. Actigraphy-measured TST exhibited a positive correlation with BMI, VAT, and BF%, while no significant correlation was found between app-measured TST and either BMI, VAT, or BF%. CONCLUSIONS: Our findings suggest that IS is strongly correlated with various adiposity indicators. Further exploration of the role of circadian rhythm, particularly measured through human-smartphone interactions, in obesity prevention could be warranted.

Developing Methods for Assessing Mental Activity Using Human-Smartphone Interactions: Comparative Analysis of Activity Levels and Phase Patterns in General Mental Activities, Working Mental Activities, and Physical Activities.

BACKGROUND: Human biological rhythms are commonly assessed through physical activity (PA) measurement, but mental activity may offer a more substantial reflection of human biological rhythms. OBJECTIVE: This study proposes a novel approach based on human-smartphone interaction to compute mental activity, encompassing general mental activity (GMA) and working mental activity (WMA). METHODS: A total of 24 health care professionals participated, wearing wrist actigraphy devices and using the "Staff Hours" app for more than 457 person-days, including 332 workdays and 125 nonworkdays. PA was measured using actigraphy, while GMA and WMA were assessed based on patterns of smartphone interactions. To model WMA, machine learning techniques such as extreme gradient boosting and convolutional neural networks were applied, using human-smartphone interaction patterns and GPS-defined work hours. The data were organized by date and divided into person-days, with an 80:20 split for training and testing data sets to minimize overfitting and maximize model robustness. The study also adopted the M10 metric to quantify daily activity levels by calculating the average acceleration during the 10-hour period of highest activity each day, which facilitated the assessment of the interrelations between PA, GMA, and WMA and sleep indicators. Phase differences, such as those between PA and GMA, were defined using a second-order Butterworth filter and Hilbert transform to extract and calculate circadian rhythms and instantaneous phases. This calculation involved subtracting the phase of the reference signal from that of the target signal and averaging these differences to provide a stable and clear measure of the phase relationship between the signals. Additionally, multilevel modeling explored associations between sleep indicators (total sleep time, midpoint of sleep) and next-day activity levels, accounting for the data's nested structure. RESULTS: Significant differences in activity levels were noted between workdays and nonworkdays, with WMA occurring approximately 1.08 hours earlier than PA during workdays (P<.001). Conversely, GMA was observed to commence about 1.22 hours later than PA (P<.001). Furthermore, a significant negative correlation was identified between the activity level of WMA and the previous night's midpoint of sleep (ß=-0.263, P<.001), indicating that later bedtimes and wake times were linked to reduced activity levels in WMA the following day. However, there was no significant correlation between WMA's activity levels and total sleep time. Similarly, no significant correlations were found between the activity levels of PA and GMA and sleep indicators from the previous night. CONCLUSIONS: This study significantly advances the understanding of human biological rhythms by developing and highlighting GMA and WMA as key indicators, derived from human-smartphone interactions. These findings offer novel insights into how mental activities, alongside PA, are intricately linked to sleep patterns, emphasizing the potential of GMA and WMA in behavioral and health studies.

Pictorial Essay on Ultrasound and Magnetic Resonance Imaging of Paraspinal Muscles for Myofascial Pain Syndrome.

The paraspinal muscles of the cervical, thoracic, and lumbar spine are important pain generators because muscle strains or myofascial pain syndrome caused by trigger points are common during clinical practice. Ultrasonography is the most convenient imaging tool for evaluating these muscles due to its advantages, such as providing good delineation of soft tissues, easy accessibility, and zero radiation. Additionally, ultrasound can serve as a useful guiding tool for paraspinal muscle intervention to prevent inadvertent injuries to vital axial neurovascular structures. This pictorial essay presents ultrasound scanning protocols for the paraspinal and other associated muscles as well as a discussion of their clinical relevance. Axial magnetic resonance imaging has also been used to elucidate reciprocal anatomy. In conclusion, ultrasound imaging proves to be a valuable tool that facilitates the differentiation of individual paraspinal muscles. This capability significantly enhances the precision of interventions designed to address myofascial pain syndrome.

Restoring functional TDP-43 oligomers in ALS and laminopathic cellular models through baicalein-induced reconfiguration of TDP-43 aggregates.

A group of misfolded prone-to-aggregate domains in disease-causing proteins has recently been shown to adopt unique conformations that play a role in fundamental biological processes. These processes include the formation of membrane-less sub-organelles, alternative splicing, and gene activation and silencing. The cellular responses are regulated by the conformational switching of prone-to-aggregate domains, independently of changes in RNA or protein expression levels. Given this, targeting the misfolded states of disease-causing proteins to redirect them towards their physiological conformations is emerging as an effective therapeutic strategy for diseases caused by protein misfolding. In our study, we successfully identified baicalein as a potent structure-correcting agent. Our findings demonstrate that baicalein can reconfigure existing TDP-43 aggregates into an oligomeric state both in vitro and in disease cells. This transformation effectively restores the bioactivity of misfolded TDP-43 proteins in cellular models of ALS and premature aging in progeria. Impressively, in progeria cells where defective lamin A interferes with TDP-43-mediated exon skipping, the formation of pathological TDP-43 aggregates is promoted. Baicalein, however, restores the functionality of TDP-43 and mitigates nuclear shape defects in these laminopathic cells. This establishes a connection between lamin A and TDP-43 in the context of aging. Our findings suggest that targeting physiological TDP-43 oligomers could offer a promising therapeutic avenue for treating aging-associated disorders.

Lidocaine inhibits migration of tenocytes by downregulating focal adhesion kinase and paxillin phosphorylation.

Lidocaine is the most frequently applied local infiltration anesthetic agent for treating tendinopathies. However, studies have discovered lidocaine to negatively affect tendon healing. In the current study, the molecular mechanisms and effects of lidocaine on tenocyte migration were evaluated. We treated tenocytes intrinsic to the Achilles tendons of Sprague-Dawley rats with lidocaine. The migration ability of cells was analyzed using electric cell-substrate impedance sensing (ECIS) and scratch wound assay. We then used a microscope to evaluate the cell spread. We assessed filamentous actin (F-actin) cytoskeleton formation through immunofluorescence staining. In addition, we used Western blot analysis to analyze the expression of phospho-focal adhesion kinase (FAK), FAK, phospho-paxillin, paxillin, and F-actin. We discovered that lidocaine had an inhibitory effect on the migration of tenocytes in the scratch wound assay and on the ECIS chip. Lidocaine treatment suppressed cell spreading and changed the cell morphology and F-actin distribution. Lidocaine reduced F-actin formation in the tenocyte during cell spreading; furthermore, it inhibited phospho-FAK, F-actin, and phospho-paxillin expression in the tenocytes. Our study revealed that lidocaine inhibits the spread and migration of tenocytes. The molecular mechanism potentially underlying this effect is downregulation of F-actin, phospho-FAK, and phospho-paxillin expression when cells are treated with lidocaine.

Overexpressing plant ferredoxin-like protein enhances photosynthetic efficiency and carbohydrates accumulation in Phalaenopsis.

Crassulacean acid metabolism (CAM) is one of three major models of carbon dioxide assimilation pathway with better water-use efficiency and slower photosynthetic efficiency in photosynthesis. Previous studies indicated that the gene of sweet pepper plant ferredoxin-like protein (PFLP) shows high homology to the ferredoxin-1(Fd-1) family that belongs to photosynthetic type Fd and involves in photosystem I. It is speculated that overexpressing pflp in the transgenic plant may enhance photosynthetic efficiency through the electron transport chain (ETC). To reveal the function of PFLP in photosynthetic efficiency, pflp transgenic Phalaenopsis, a CAM plant, was generated to analyze photosynthetic markers. Transgenic plants exhibited 1.2-folds of electron transport rate than that of wild type (WT), and higher CO2 assimilation rates up to 1.6 and 1.5-folds samples at 4 pm and 10 pm respectively. Enzyme activity of phosphoenolpyruvate carboxylase (PEPC) was increased to 5.9-folds in Phase III, and NAD+-linked malic enzyme (NAD+-ME) activity increased 1.4-folds in Phase IV in transgenic plants. The photosynthesis products were analyzed between transgenic plants and WT. Soluble sugars contents such as glucose, fructose, and sucrose were found to significantly increase to 1.2, 1.8, and 1.3-folds higher in transgenic plants. The starch grains were also accumulated up to 1.4-folds in transgenic plants than that of WT. These results indicated that overexpressing pflp in transgenic plants increases carbohydrates accumulation by enhancing electron transport flow during photosynthesis. This is the first evidence for the PFLP function in CAM plants. Taken altogether, we suggest that pflp is an applicable gene for agriculture application that enhances electron transport chain efficiency during photosynthesis.

DNA Damage and Nuclear Morphological Changes in Cardiac Hypertrophy Are Mediated by SNRK Through Actin Depolymerization.

BACKGROUND: Proper nuclear organization is critical for cardiomyocyte function, because global structural remodeling of nuclear morphology and chromatin structure underpins the development and progression of cardiovascular disease. Previous reports have implicated a role for DNA damage in cardiac hypertrophy; however, the mechanism for this process is not well delineated. AMPK (AMP-activated protein kinase) family of proteins regulates metabolism and DNA damage response (DDR). Here, we examine whether a member of this family, SNRK (SNF1-related kinase), which plays a role in cardiac metabolism, is also involved in hypertrophic remodeling through changes in DDR and structural properties of the nucleus. METHODS: We subjected cardiac-specific Snrk-/- mice to transaortic banding to assess the effect on cardiac function and DDR. In parallel, we modulated SNRK in vitro and assessed its effects on DDR and nuclear parameters. We also used phosphoproteomics to identify novel proteins that are phosphorylated by SNRK. Last, coimmunoprecipitation was used to verify Destrin (DSTN) as the binding partner of SNRK that modulates its effects on the nucleus and DDR. RESULTS: Cardiac-specific Snrk-/- mice display worse cardiac function and cardiac hypertrophy in response to transaortic banding, and an increase in DDR marker pH2AX (phospho-histone 2AX) in their hearts. In addition, in vitro Snrk knockdown results in increased DNA damage and chromatin compaction, along with alterations in nuclear flatness and 3-dimensional volume. Phosphoproteomic studies identified a novel SNRK target, DSTN, a member of F-actin depolymerizing factor proteins that directly bind to and depolymerize F-actin. SNRK binds to DSTN, and DSTN downregulation reverses excess DNA damage and changes in nuclear parameters, in addition to cellular hypertrophy, with SNRK knockdown. We also demonstrate that SNRK knockdown promotes excessive actin depolymerization, measured by the increased ratio of G-actin to F-actin. Last, jasplakinolide, a pharmacological stabilizer of F-actin, rescues the increased DNA damage and aberrant nuclear morphology in SNRK-downregulated cells. CONCLUSIONS: These results indicate that SNRK is a key player in cardiac hypertrophy and DNA damage through its interaction with DSTN. This interaction fine-tunes actin polymerization to reduce DDR and maintain proper cardiomyocyte nuclear shape and morphology.

Measurement of Thermal Stress by X-ray Nano-Diffraction in (111)-Oriented Nanotwinned Cu Bumps for Cu/SiO2 Hybrid Joints.

X-ray nanodiffraction was used to measure the thermal stress of 10 µm nanotwinned Cu bumps in Cu/SiO2 hybrid structures at -55 °C, 27 °C, 100 °C, 150 °C, and 200 °C. Bonding can be achieved without externally applied compression. The X-ray beam size is about 100 nm in diameter. The Cu bump is dominated by (111) oriented nano-twins. Before the hybrid bonding, the thermal stress in Cu bumps is compressive and remains compressive after bonding. The average stress in the bonded Cu joint at 200 °C is as large as -169.1 MPa. In addition, using the strain data measured at various temperatures, one can calculate the effective thermal expansion coefficient (CTE) for the 10 µm Cu bumps confined by the SiO2 dielectrics. This study reports a useful approach on measuring the strain and stress in oriented metal bumps confined by SiO2 dielectrics. The results also provide a deeper understanding on the mechanism of hybrid bonding without externally applied compression.

Iron drives anabolic metabolism through active histone demethylation and mTORC1.

All eukaryotic cells require a minimal iron threshold to sustain anabolic metabolism. However, the mechanisms by which cells sense iron to regulate anabolic processes are unclear. Here we report a previously undescribed eukaryotic pathway for iron sensing in which molecular iron is required to sustain active histone demethylation and maintain the expression of critical components of the pro-anabolic mTORC1 pathway. Specifically, we identify the iron-binding histone-demethylase KDM3B as an intrinsic iron sensor that regulates mTORC1 activity by demethylating H3K9me2 at enhancers of a high-affinity leucine transporter, LAT3, and RPTOR. By directly suppressing leucine availability and RAPTOR levels, iron deficiency supersedes other nutrient inputs into mTORC1. This process occurs in vivo and is not an indirect effect by canonical iron-utilizing pathways. Because ancestral eukaryotes share homologues of KDMs and mTORC1 core components, this pathway probably pre-dated the emergence of the other kingdom-specific nutrient sensors for mTORC1.

Simultaneous enhancement of the bandwidth and responsivity in high-speed avalanche photodiodes with an optimized flip-chip bonding package.

The enhancement in responsivity of photodiodes (PDs) or avalanche photodiodes (APDs) with the traditional flip-chip bonding package usually comes at the expense of degradation in the optical-to-electrical (O-E) bandwidth due to the increase of parasitic capacitance. In this work, we demonstrate backside-illuminated In0.52Al0.48As based APDs with novel flip-chip bonding packaging designed to relax this fundamental trade-off. The inductance induced peak in the measured O-E frequency response of these well-designed and well-packaged APDs, which can be observed around its 3-dB bandwidth (∼30 GHz), effectively widens the bandwidth and becomes more pronounced when the active diameter of the APD is aggressively downscaled to as small as 3 µm. With a typical active window diameter of 14 µm, large enough for alignment tolerance and low optical coupling loss, the packaged APD exhibits a moderate damping O-E frequency response with a bandwidth (36 vs. 31 GHz) and responsivity (3.4 vs. 2.3 A/W) superior to those of top-illuminated reference sample under 0.9 Vbr operation, to attain a high millimeter wave output power (0 dBm at 40 GHz) and output current (12.5 mA at +8.8 dBm optical power). The excellent static and dynamic performance of this design open up new possibilities to further improve the sensitivity at the receiver-end of the next-generation of passive optical network (PON) and coherent communication systems.

SIRT2 inhibition protects against cardiac hypertrophy and ischemic injury.

Sirtuins (SIRT) exhibit deacetylation or ADP-ribosyltransferase activity and regulate a wide range of cellular processes in the nucleus, mitochondria, and cytoplasm. The role of the only sirtuin that resides in the cytoplasm, SIRT2, in the development of ischemic injury and cardiac hypertrophy is not known. In this paper, we show that the hearts of mice with deletion of Sirt2 (Sirt2-/-) display improved cardiac function after ischemia-reperfusion (I/R) and pressure overload (PO), suggesting that SIRT2 exerts maladaptive effects in the heart in response to stress. Similar results were obtained in mice with cardiomyocyte-specific Sirt2 deletion. Mechanistic studies suggest that SIRT2 modulates cellular levels and activity of nuclear factor (erythroid-derived 2)-like 2 (NRF2), which results in reduced expression of antioxidant proteins. Deletion of Nrf2 in the hearts of Sirt2-/- mice reversed protection after PO. Finally, treatment of mouse hearts with a specific SIRT2 inhibitor reduced cardiac size and attenuates cardiac hypertrophy in response to PO. These data indicate that SIRT2 has detrimental effects in the heart and plays a role in cardiac response to injury and the progression of cardiac hypertrophy, which makes this protein a unique member of the SIRT family. Additionally, our studies provide a novel approach for treatment of cardiac hypertrophy and injury by targeting SIRT2 pharmacologically, providing a novel avenue for the treatment of these disorders.

Structural basis for calcium-stimulating pore formation of Vibrio α-hemolysin.

Vibrio α-hemolysins (αHLs) are ß-pore-forming toxins secreted by Vibrio pathogens, crucial for the facilitation of bacterial infections through host cell lysis. These toxins are produced as inactive precursors, requiring proteolytic maturation and membrane association for activation within host tissues. Here, we investigate Vibrio campbellii αHL (VcαHL), and establish that its hemolytic activity is significantly stimulated by calcium ions, with an EC50 that aligns with physiological calcium concentrations. Furthermore, we illustrate the vital contribution of calcium ions to the oligomerization of VcαHL on membranes. Using X-ray crystallography and cryo-electron microscopy, we decipher both the immature and assembled structures of VcαHL and elucidate the conformational changes corresponding to toxin assembly. We also identify a calcium-binding module that is integral for VcαHL's calcium-dependent activation. These findings provide insights into the regulatory mechanisms of VcαHL and have the potential to inform the development of targeted therapeutic strategies against Vibrio infections.

Indoor CO2 monitoring in a surgical intensive care unit under visitation restrictions during the COVID-19 pandemic.

Background: Indoor CO2 concentration is an important metric of indoor air quality (IAQ). The dynamic temporal pattern of CO2 levels in intensive care units (ICUs), where healthcare providers experience high cognitive load and occupant numbers are frequently changing, has not been comprehensively characterized. Objective: We attempted to describe the dynamic change in CO2 levels in the ICU using an Internet of Things-based (IoT-based) monitoring system. Specifically, given that the COVID-19 pandemic makes hospital visitation restrictions necessary worldwide, this study aimed to appraise the impact of visitation restrictions on CO2 levels in the ICU. Methods: Since February 2020, an IoT-based intelligent indoor environment monitoring system has been implemented in a 24-bed university hospital ICU, which is symmetrically divided into areas A and B. One sensor was placed at the workstation of each area for continuous monitoring. The data of CO2 and other pollutants (e.g., PM2.5) measured under standard and restricted visitation policies during the COVID-19 pandemic were retrieved for analysis. Additionally, the CO2 levels were compared between workdays and non-working days and between areas A and B. Results: The median CO2 level (interquartile range [IQR]) was 616 (524-682) ppm, and only 979 (0.34%) data points obtained in area A during standard visitation were ≥ 1,000 ppm. The CO2 concentrations were significantly lower during restricted visitation (median [IQR]: 576 [556-596] ppm) than during standard visitation (628 [602-663] ppm; p < 0.001). The PM2.5 concentrations were significantly lower during restricted visitation (median [IQR]: 1 [0-1] µg/m3) than during standard visitation (2 [1-3] µg/m3; p < 0.001). The daily CO2 and PM2.5 levels were relatively low at night and elevated as the occupant number increased during clinical handover and visitation. The CO2 concentrations were significantly higher in area A (median [IQR]: 681 [653-712] ppm) than in area B (524 [504-547] ppm; p < 0.001). The CO2 concentrations were significantly lower on non-working days (median [IQR]: 606 [587-671] ppm) than on workdays (583 [573-600] ppm; p < 0.001). Conclusion: Our study suggests that visitation restrictions during the COVID-19 pandemic may affect CO2 levels in the ICU. Implantation of the IoT-based IAQ sensing network system may facilitate the monitoring of indoor CO2 levels.

DNA damage and nuclear morphological changes in cardiac hypertrophy are mediated by SNRK through actin depolymerization.

BACKGROUND: Proper nuclear organization is critical for cardiomyocyte (CM) function, as global structural remodeling of nuclear morphology and chromatin structure underpins the development and progression of cardiovascular disease. Previous reports have implicated a role for DNA damage in cardiac hypertrophy, however, the mechanism for this process is not well delineated. AMPK family of proteins regulate metabolism and DNA damage response (DDR). Here, we examine whether a member of this family, SNF1-related kinase (SNRK), which plays a role in cardiac metabolism, is also involved in hypertrophic remodeling through changes in DDR and structural properties of the nucleus. METHODS: We subjected cardiac specific (cs)- Snrk -/- mice to trans-aortic banding (TAC) to assess the effect on cardiac function and DDR. In parallel, we modulated SNRK in vitro and assessed its effects on DDR and nuclear parameters. We also used phospho-proteomics to identify novel proteins that are phosphorylated by SNRK. Finally, co-immunoprecipitation (co-IP) was used to verify Destrin (DSTN) as the binding partner of SNRK that modulates its effects on the nucleus and DDR. RESULTS: cs- Snrk -/- mice display worse cardiac function and cardiac hypertrophy in response to TAC, and an increase in DDR marker pH2AX in their hearts. Additionally, in vitro Snrk knockdown results in increased DNA damage and chromatin compaction, along with alterations in nuclear flatness and 3D volume. Phospho-proteomic studies identified a novel SNRK target, DSTN, a member of F-actin depolymerizing factor (ADF) proteins that directly binds to and depolymerize F-actin. SNRK binds to DSTN, and DSTN downregulation reverses excess DNA damage and changes in nuclear parameters, in addition to cellular hypertrophy, with SNRK knockdown. We also demonstrate that SNRK knockdown promotes excessive actin depolymerization, measured by the increased ratio of globular (G-) actin to F-actin. Finally, Jasplakinolide, a pharmacological stabilizer of F-actin, rescues the increased DNA damage and aberrant nuclear morphology in SNRK downregulated cells. CONCLUSIONS: These results indicate that SNRK is a key player in cardiac hypertrophy and DNA damage through its interaction with DSTN. This interaction fine-tunes actin polymerization to reduce DDR and maintain proper CM nuclear shape and morphology. Clinical Perspective: What is new? Animal hearts subjected to pressure overload display increased SNF1-related kinase (SNRK) protein expression levels and cardiomyocyte specific SNRK deletion leads to aggravated myocardial hypertrophy and heart failure.We have found that downregulation of SNRK impairs DSTN-mediated actin polymerization, leading to maladaptive changes in nuclear morphology, higher DNA damage response (DDR) and increased hypertrophy. What are the clinical implications? Our results suggest that disruption of DDR through genetic loss of SNRK results in an exaggerated pressure overload-induced cardiomyocyte hypertrophy.Targeting DDR, actin polymerization or SNRK/DSTN interaction represent promising therapeutic targets in pressure overload cardiac hypertrophy.

Synergistic Chemopreventive Effects of a Novel Combined Plant Extract Comprising Gallic Acid and Hesperidin on Colorectal Cancer.

BACKGROUND/AIM: Colorectal cancer (CRC) is the third most common cancer with a high mortality rate worldwide. Although gallic acid and hesperidin exert anticancer activity, synergistic effects of gallic acid and hesperidin against CRC remain elusive. This study aims to investigate the therapeutic mechanism of a novel combination of gallic acid and hesperidin against CRC cell growth, including cell viability, cell-cycle-associated proteins, spheroid formation, and stemness. METHODS: Gallic acid and hesperidin derived from Hakka pomelo tea (HPT) were detected by colorimetric methods and high-performance liquid chromatography using ethyl acetate as an extraction medium. CRC cell lines (HT-29 and HCT-116) treated with the combined extract were investigated in our study for cell viability (trypan blue or soft agar colony formation assay), cell cycle (propidium iodide staining), cell-cycle-associated proteins (immunoblotting), and stem cell markers (immunohistochemistry staining). RESULTS: Compared with other extraction methods, HPT extraction using an ethyl acetate medium exerts the most potent effect on inhibiting HT-29 cell growth in a dose-dependent manner. Furthermore, the treatment with combined extract had a higher inhibitory effect on CRC cell viability than gallic acid or hesperidin alone. The underlying mechanism was involved in G1-phase arrest and Cip1/p21 upregulation that could attenuate HCT-116 cell proliferation (Ki-67), stemness (CD-133), and spheroid growth in a 3D formation assay mimicking in vivo tumorigenesis. CONCLUSION: Gallic acid and hesperidin exert synergistic effects on cell growth, spheroids, and stemness of CRC and may serve as a potential chemopreventive agent. Further testing for the safety and effectiveness of the combined extract in large-scale randomized trials is required.

SIRT2 inhibition protects against cardiac hypertrophy and heart failure.

Sirtuins (SIRT) exhibit deacetylation or ADP-ribosyltransferase activity and regulate a wide range of cellular processes in the nucleus, mitochondria and cytoplasm. The role of the only sirtuin that resides in the cytoplasm, SIRT2, in the development of heart failure (HF) and cardiac hypertrophy is not known. In this paper, we show that the hearts of mice with deletion of Sirt2 ( Sirt2 -/- ) display improved cardiac function after ischemia-reperfusion (I/R) and pressure overload (PO), suggesting that SIRT2 exerts maladaptive effects in the heart in response to stress. Similar results were obtained in mice with cardiomyocyte-specific Sirt2 deletion. Mechanistic studies suggest that SIRT2 modulates cellular levels and activity of nuclear factor (erythroid-derived 2)-like 2 (NRF2), which results in reduced expression of antioxidant proteins. Deletion of Nrf2 in the hearts of Sirt2 -/- mice reversed protection after PO. Finally, treatment of mouse hearts with a specific SIRT2 inhibitors reduces cardiac size and attenuates cardiac hypertrophy in response to PO. These data indicate that SIRT2 has detrimental effects in the heart and plays a role in the progression of HF and cardiac hypertrophy, which makes this protein a unique member of the SIRT family. Additionally, our studies provide a novel approach for treatment of cardiac hypertrophy by targeting SIRT2 pharmacologically, providing a novel avenue for the treatment of this disorder.

Optimized protocol for quantification of mitochondrial non-heme and heme iron content in mouse tissues and cultured cells.

Impaired mitochondrial iron metabolism is associated with aging and a variety of diseases, and there is a growing need to accurately quantify mitochondrial iron levels. This protocol provides an optimized method for evaluating non-heme and heme iron in mitochondrial and cytosolic fractions of tissues and cultured cells. Our protocol consists of three steps: sample fractionation, non-heme iron measurement, and heme iron measurement. For complete details on the use and execution of this protocol, please refer to Sato et al. (2022).1.