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Abstract Objectives To systematically evaluate the therapeutic effect of electrical stimulation combined with pelvic floor muscle exercise on female pelvic floor dysfunction (PFD). Methods Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) was applied. A computer-based retrieval was performed in the databases of PubMed, Web of Science, Embase, and Cochrane Library from database establishment to September 15, 2023, to identify randomized controlled trials on electrical stimulation combined with pelvic floor muscle function exercise on female PFD. Literature screening, data extraction, and quality evaluation were performed independently by two researchers, and meta-analysis was performed using the statistical software Stata15.0. Results 1. In total, 12 randomized controlled trials were included, involving 721 female patients. The overall quality of methodologies employed in the included studies was relatively high. 2. Meta-analysis results showed that electrical stimulation combined with pelvic floor muscle exercise could effectively mitigate the severity of female PFD (SMD = -1.01, 95% CI − 1.78, − 0.25, P 0.05). Conclusions Compared with pelvic floor muscle exercise alone, electrical stimulation combined with pelvic floor muscle exercise could effectively mitigate the severity of female PFD. It had a notable positive impact on enhancing pelvic floor muscle strength in female patients, although it did not significantly improve quality of life. Future high-quality studies are warranted.

Abstract Introduction The prognostic value of multifocality in paediatric papillary thyroid carcinoma (PTC) patients remains a subject of debate. This study aimed to explore the clinical significance and prognostic value of multifocality in children and adolescents with PTC. Methods This study retrospectively analysed the clinicopathological characteristics and postoperative follow-up data of 338 PTC patients aged ≤ 20 years from May 2012 to July 2022. The clinical and pathological characteristics of 205 patients with unifocal lesions and 133 patients with multifocal lesions were compared. A logistic regression model evaluated the relationship between multifocal lesions and disease recurrence/persistence in children and adolescents with PTC. Based on the median follow-up time of children with multifocal PTC, 114 patients with multifocal PTC older than 20 years were added, and the clinicopathological characteristics were compared between the 133. paediatric/adolescent patients and 114 adult patients with multifocal PTC. Results Among the paediatric and adolescent patients, over a median follow-up time of 49 months, 133 had multifocal disease and 205 had unifocal disease. Multifocal PTC patients exhibited stronger invasiveness in the form of extrathyroidal extension, tumour diameter, lymph node metastasis, and distant metastasis. Multifocality (OR 2.68; p = 0.017), lateral lymph node metastasis (OR 2.85; p = 0.036), and distant metastasis (OR 4.28; p = 0.010) were identified as independent predictive factors for the recurrence/persistence of disease. Comparing the paediatric/adolescent vs. adult multifocal patients, the former demonstrated greater tumour invasiveness. Lateral lymph node metastasis (OR 6.36; P = 0.012) and distant metastasis (OR 3.70; P = 0.027) were independent predictive factors for recurrence/persistence of disease in multifocal patients, while age was not (OR 0.95; P = 0.455). Conclusion Tumour multifocality independently predicts persistent/recurrent disease in paediatric and adolescent PTC patients.

IntroductionThyroid cancer (THCA) is the most common endocrine tumor. Coagulation may be associated with the development of cancer, but its role in THCA patients is not yet clear.MethodsIn this study, we determined the predictive value of coagulation biomarker D-dimer for THCA patient lateral lymph node metastasis (LLNM) through receiver operating characteristics (ROC) analysis and logistic regression analysis. Subsequently, this study used the TCGA database to identify coagulation-related molecular subtypes through consensus clustering analysis and compared their prognosis. We identified coagulation-related genes (CRGs) associated with prognosis in thyroid cancer through gene expression data and clinical information, and constructed a prognostic model by selecting the prognostic CRGs using LASSO regression. Patients were divided into high-risk and low-risk groups based on the median score. Subsequently, prognosis, clinical characteristics, gene mutation occurrence, immune infiltration, function, and drug sensitivity of the two groups were analyzed. We also constructed a nomogram combining the model and clinical features. Finally, the expression of the prognostic CRGs was validated by RT-qPCR.ResultsD-dimers had better performance in predicting LLNM(the area under the curve was 0.656 (95% CI 0.580-0.733), with a cut-off value of 0.065 mg/l), and D-dimer>0.065mg/l was an independent predictor of LLNM. Then, we selected 8 prognostic CRGs to construct a predictive model. The prognosis of low-risk group patients was significantly better than that of high-risk group (P

Abstract While hypoxic signaling has been shown to play a role in many cellular processes, its role in metabolism-linked extracellular matrix (ECM) organization and downstream processes of cell fate after musculoskeletal injury remains to be determined. Heterotopic ossification (HO) is a debilitating condition where abnormal bone formation occurs within extra-skeletal tissues. Hypoxia and hypoxia-inducible factor 1α (HIF-1α) activation have been shown to promote HO. However, the underlying molecular mechanisms by which the HIF-1α pathway in mesenchymal progenitor cells (MPCs) contributes to pathologic bone formation remain to be elucidated. Here, we used a proven mouse injury-induced HO model to investigate the role of HIF-1α on aberrant cell fate. Using single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics analyses of the HO site, we found that collagen ECM organization is the most highly up-regulated biological process in MPCs. Zeugopod mesenchymal cell-specific deletion of Hif1α (Hoxa11-CreER T2 ; Hif1a fl/fl ) significantly mitigated HO in vivo. ScRNA-seq analysis of these Hoxa11-CreER T2 ; Hif1a fl/fl mice identified the PLOD2/LOX pathway for collagen cross-linking as downstream of the HIF-1α regulation of HO. Importantly, our scRNA-seq data and mechanistic studies further uncovered that glucose metabolism in MPCs is most highly impacted by HIF-1α deletion. From a translational aspect, a pan-LOX inhibitor significantly decreased HO. A newly screened compound revealed that the inhibition of PLOD2 activity in MPCs significantly decreased osteogenic differentiation and glycolytic metabolism. This suggests that the HIF-1α/PLOD2/LOX axis linked to metabolism regulates HO-forming MPC fate. These results suggest that the HIF-1α/PLOD2/LOX pathway represents a promising strategy to mitigate HO formation.

Abstract The live transport of farmed fish is an important practice in Chinese aquaculture due to consumer preferences in its domestic market. However, live transport can be stressful for fish and may cause many welfare issues. This study aimed to examine the effects of transport density on the welfare of largemouth bass (Micropterus salmoides). Adult fish were allocated to four experimental treatments in a two‐factor design (N = 5 groups per treatment). Factor one was stocking density of either 39 (low density) or 78 (high density) g L−1. Factor two was transport, with half of the fish groups exposed to simulated transport stress for 3 h using a motion platform, and the other half kept in static tanks for the same length of time. Baseline behavior and physiology were determined from a subset of fish after acclimation but before selection into groups and application of transport. To determine the effects of transport, stocking density, and their interaction on the experimental groups, physiological sampling and behavioral recording were carried out immediately after simulated transport, and water quality was measured both before and after transport. Changes in water quality were observed between two timepoints, with decreased dissolved oxygen levels and increased water temperature. Total ammonia nitrogen levels significantly increased in all treatments over time, particularly in the high‐density groups. Fish ventilation rates were significantly elevated in the groups kept at a high stocking density regardless of whether they also experienced transport stress. Fish transported at a high stocking density also showed more fear‐related behaviors such as freezing and thigmotaxis than non‐transported groups. Serum cortisol, catalase, and aspartate transaminase levels were significantly higher in the transported treatment groups compared to the baseline fish, but no difference was found for serum glucose, lactate, and glutathione peroxidase. We concluded that 3 h of simulated transport was stressful to largemouth bass kept at 78 g L−1, as evidenced by altered fish physiological and behavioral indicators of stress, as well as deteriorated water quality. Additionally, we identified a positive correlation between serum and skin mucus samples for cortisol, glucose, and lactate in transported fish, providing important information for developing less invasive methods for the welfare assessment of largemouth bass.

Summary: Heterotopic ossification (HO) is a challenging condition that occurs after musculoskeletal injury and is characterized by the formation of bone in non-skeletal tissues. While the effect of HO on blood vessels is well established, little is known about its impact on lymphatic vessels. Here, we use a mouse model of traumatic HO to investigate the relationship between HO and lymphatic vessels. We show that injury triggers lymphangiogenesis at the injury site, which is associated with elevated vascular endothelial growth factor C (VEGF-C) levels. Through single-cell transcriptomic analyses, we identify mesenchymal progenitor cells and tenocytes as sources of Vegfc. We demonstrate by lineage tracing that Vegfc-expressing cells undergo osteochondral differentiation and contribute to the formation of HO. Last, we show that Vegfc haploinsufficiency results in a nearly 50% reduction in lymphangiogenesis and HO formation. These findings shed light on the complex mechanisms underlying HO formation and its impact on lymphatic vessels.

Abstract Ionizing radiation (IR) causes a wide variety of DNA lesions, of which DNA double-stranded breaks (DSBs) are the most deleterious. Homologous recombination (HR) is a crucial route responsible for repairing DSBs. RecQ-mediated genome instability protein 1 (RMI1) is a member of an evolutionarily conserved Bloom syndrome complex, which prevents and resolves aberrant recombination products during HR, thereby promoting genome stability. However, little is known about the role of RMI1 in regulating the cellular response to IR. This study aimed to understand the cellular functions and molecular mechanisms by which RMI1 maintains genomic stability after IR exposure. Here, we showed IR upregulated the RMI1 protein level and induced RMI1 relocation to the DNA damage sites. We also demonstrated that the loss of RMI1 in cells resulted in enhanced levels of DNA damage, sustained cell cycle arrest, and impaired HR repair after IR, leading to reduced cell viability and elevated genome instability. Taken together, our results highlighted the direct roles of RMI1 in response to DNA damage induced by IR and implied that RMI1 might be a new genome safeguard molecule to radiation-induced damage.

Abstract Background The transmembrane receptor Kremen2 has been reported to participate in the tumorigenesis and metastasis of gastric cancer. However, the role of Kremen2 in non-small cell lung cancer (NSCLC) and the underlying mechanism remain unclear. This study aimed to explore the biological function and regulatory mechanism of Kremen2 in NSCLC. Methods The correlation between Kremen2 expression and NSCLC was assessed by analyzing the public database and clinical tissue samples. Colony formation and EdU assays were performed to examine cell proliferation. Transwell and wound healing assays were used to observe cell migration ability. Tumor-bearing nude mice and metastatic tumor models were used to detect the in vivo tumorigenic and metastatic abilities of the NSCLC cells. An immunohistochemical assay was used to detect the expression of proliferation-related proteins in tissues. Western blot, immunoprecipitation and immunofluorescence were conducted to elucidate the Kremen2 regulatory mechanisms in NSCLC. Results Kremen2 was highly expressed in tumor tissues from NSCLC patients and was positively correlated with a poor patient prognosis. Knockout or knockdown of Kremen2 inhibited cell proliferation and migration ability of NSCLC cells. In vivo knockdown of Kremen2 inhibited the tumorigenicity and number of metastatic nodules of NSCLC cells in nude mice. Mechanistically, Kremen2 interacted with suppressor of cytokine signaling 3 (SOCS3) to maintain the epidermal growth factor receptor (EGFR) protein levels by preventing SOCS3-mediated ubiquitination and degradation of EGFR, which, in turn, promoted activation of the PI3K-AKT and JAK2-STAT3 signaling pathways. Conclusions Our study identified Kremen2 as a candidate oncogene in NSCLC and may provide a potential target for NSCLC treatment.

Abstract The functional interdependence of nerves and blood vessels is a well-established concept during tissue morphogenesis, yet the role of neurovascular coupling in proper and aberrant tissue repair is an emerging field of interest. Here, we sought to define the regulatory relationship of peripheral nerves on vasculature in a severe extremity trauma model in mice, which results in aberrant cell fate and heterotopic ossification (HO). First, a high spatial degree of neurovascular congruency was observed to exist within extremity injury associated heterotopic ossification. Vascular and perivascular cells demonstrate characteristic responses to injury, as assessed by single cell RNA sequencing. This vascular response to injury was blunted in neurectomized mice, including a decrease in endothelial proliferation and type H vessel formation, and a downregulation of key transcriptional networks associated with angiogenesis. Independent mechanisms to chemically or genetically inhibit axonal ingrowth led to similar deficits in HO site angiogenesis, a reduction in type H vessels, and heterotopic bone formation. Finally, a combination of single cell transcriptomic approaches within the dorsal root ganglia identified key neural-derived angiogenic paracrine factors that may mediate neuron-to-vascular signaling in HO. These data provide further understanding of nerve-to-vessel crosstalk in traumatized soft tissues, which may reflect a key determinant of mesenchymal progenitor cell fate after injury.

Gaucher disease (GD) is an inherited lysosomal storage disease caused by mutations in the glucocerebrosidase gene. The decrease of glucocerebrosidase activity in lysosomes results in the accumulation of its substrate glucocerebroside in the lysosomes of macrophages in organs such as the liver, spleen, bones, lungs, brain and eyes, and the formation of typical storage cells, namely “Gaucher cells”, leading to lesions in the affected tissues and organs. Hepatosplenomegaly, bone pain, cytopenia, neurological symptoms, and other systemic manifestations are common in clinical practice. Most pediatric patients have severe symptoms. Early diagnosis and treatment are crucial to improve the curative effect and prognosis. However, due to the low incidence of this disease, multi-system involvement in patients, and diverse clinical manifestations, multidisciplinary teamwork is needed for comprehensive evaluation, diagnosis and treatment. In this study, we reported 2 cases of different types of GD who were diagnosed, treated and followed up by multidisciplinary collaboration in infancy.

Background Eighty-eight percent of the persons with bilateral vestibular dysfunction have reported at least one fall within the past 5 years. The apparent alternations due to the bilateral vestibular dysfunctions (BVD) are the gait characteristics, such as slower walking speed, prolonged stance phase, and shorter step length. Unexpectedly, due to the prevalence of this BVD being relatively low, attention is not obtained as same as in other vestibular disorders. Moreover, how does walking on different inclines, part of daily activities, alter the gait characteristics under the unreliable bilateral vestibular systems? Previous studies used vibration-based stimulations (VS) as a perturbation to understand the postural control during walking while the bilateral vestibular systems were perturbed. Therefore, this study attempted to extend the knowledge to understand the alternations in spatial-temporal gait characteristics under perturbed bilateral vestibular systems while walking on different inclines. Methods Nineteen healthy young adults participated in this study. Eight walking conditions were randomly assigned to each participant: 0%, 3%, 6%, and 9% grade of inclines with/without VS. The preferred walking speed was used for gait analysis. The dependent variables were stance time, double support time, step length, step time, step width, foot clearance, and respective variabilities. All dependent variables were defined by two critical gait events: heel-strike and toe-off. Pre-Hoc paired comparisons with Bonferroni corrections were used to prioritize the dependent variables. A two-way repeated measure was used to investigate the effect of VS and the effect of inclines on the selected dependent variables from Pre-Hoc analysis. Post-Hoc comparisons were also corrected by the Bonferroni method. Results The step length, step time, foot clearance, and foot clearance variability were selected by the Pre-Hoc analysis because the corrected paired t-test demonstrated a significant VS effect (p < 0.05) on these gait parameters at least one of four inclines. The significant interaction between the effect of VS and the effect of inclines was found in step length (p = 0.005), step time (p = 0.028), and foot clearance variability (p = 0.003). The results revealed that implementing a VS increased step length and step time when walking on 0%, 3%, and 9% of grade inclines. In particular, the foot clearance variability was found when walking on 9% of grade inclines. Conclusion The observations in the current study suggested that VS increased the step length, step time, foot clearance, and foot clearance variability while walking on inclines. These results suggested that these gait parameters might be promising targets for future clinical investigations in patients with BVD while walking on different inclines. Importantly, the increases in spatial-temporal gait performance under bilateral VS might be an indicator of gait improvement while walking on different inclines.

Soft tissue trauma can result in aberrant osteochondral differentiation of local mesenchymal progenitor cells. Here the authors show that, in mice, soft tissue trauma results in NGF expression by perivascular cells, which leads to axonal invasion and drives abnormal osteochondral differentiation, and show that this process can be prevented by inhibition of NGF signaling.

Background We showed that Beclin‐1‐dependent autophagy protects the heart in young and adult mice that underwent endotoxemia. Herein, we compared the potential therapeutic effects of Beclin‐1 activating peptide, TB‐peptide, on endotoxemia‐induced cardiac outcomes in young adult and aged mice. We further evaluated lipopolysaccharide (lipopolysaccharide)‐induced and TB‐peptide treatment‐mediated alterations in myocardial metabolism. Methods and Results C57BL/6J mice that were 10 weeks and 24 months old were challenged by lipopolysaccharide using doses at which cardiac dysfunction occurred. Following the treatment of TB‐peptide or control vehicle, heart contractility, circulating cytokines, and myocardial autophagy were evaluated. We detected that TB‐peptide boosted autophagy, attenuated cytokines, and improved cardiac performance in both young and aged mice during endotoxemia. A targeted metabolomics assay was designed to detect a pool of 361 known metabolites, of which 156 were detected in at least 1 of the heart tissue samples. Lipopolysaccharide‐induced impairments were found in glucose and amino acid metabolisms in mice of all ages, and TB‐peptide ameliorated these alterations. However, lipid metabolites were upregulated in the young group but moderately downregulated in the aged by lipopolysaccharide, suggesting an age‐dependent response. TB‐peptide mitigated lipopolysaccharide‐mediated trend of lipids in the young mice but had little effect on the aged. (Study registration: Project DOI: https://doi.org/10.21228/M8K11W). Conclusions Pharmacological activation of Beclin‐1 by TB‐peptide is cardiac protective in both young and aged population during endotoxemia, suggest a therapeutic potential for sepsis‐induced cardiomyopathy. Metabolomics analysis suggests that an age‐independent protection by TB‐peptide is associated with reprograming of energy production via glucose and amino acid metabolisms.

Abstract Mitochondria‐associated membranes (MAMs) are essential to mitochondria. This study was to determine whether endotoxemia rearranges MAMs in the heart, and whether Beclin‐1 regulates this process. Wild‐type mice and mice with a cardiac‐specific overexpression of Beclin‐1 (Becn1‐Tg), or a heterozygous knockout of Beclin‐1 (Becn1+/−) were given lipopolysaccharide (LPS) challenge. In the heart, the ultrastructure of MAMs was examined by electron microscopy and the histology evaluated by immunostaining. Additionally, MAMs were isolated by ultracentrifugation, and their content and function were quantified. The effects of Beclin‐1‐activating peptide (TB‐peptide) on MAMs were also examined. Data showed that endotoxemia decreased both the total mass and the function of MAMs, and these deficiencies became worse in Becn1+/− mice but were alleviated in Becn1‐Tg and TB‐peptide‐treated mice. Responses of myocardial MAMs to LPS and to TB‐peptide were additionally examined in AC16 human cardiomyocytes. In vitro findings recaptured the effects of LPS and TB‐peptide in cardiomyocytes; the challenge of LPS reduced the level and activity of MAMs, and TB‐peptide attenuated this defect. Together, the results suggest a new function of Beclin‐1 in improving cardiac MAMs during endotoxemia, providing a mechanism for the previously identified role of Beclin‐1 in protection of mitochondria and cardiac function.

The novel coronavirus pandemic has led to morbidity and mortality throughout the world. Until now, it is a highly virulent contagion attacking the respiratory system in humans, especially people with chronic diseases and the elderly who are most vulnerable. A majority of afflicted are those suffering from cardiovascular and coronary diseases. In this review article, an attempt has been made to discuss and thoroughly review the mode of therapies that alleviate cardiac complications and complications due to hypercoagulation in patients infected with the SARS-CoV-2 virus. Presently a host of thrombolytic drugs are in use like Prourokinase, Retelapse, RhTNK-tPA and Urokinase. However, thrombolytic therapy, especially if given intravenously, is associated with a serious risk of intracranial haemorrhage, systemic haemorrhage, immunologic complications, hypotension and myocardial rupture. The effects of the SARS-CoV-2 virus upon the cardiovascular system and coagulation state of the body are being closely studied. In connection to the same, clinical prognosis and complications of thrombolytic therapy are being scrutinized. It is noteworthy to mention that myocardial oxygen supply/demand mismatch, direct myocardial cells injury and acute plaque rupture are the multiple mechanisms responsible for acute coronary syndrome and cardiac complications in Covid-19 infection. However, this review has limitations as data available in this context is limited, scattered and heterogenous that questions the reliability of the same. So, more multi-centric studies involving representative populations, carried out meticulously, could further assist in responding better to cardiac complications among Covid-19 patients.

Hypoxia-ischemia (HI) is the most common acute brain threat in neonates and a leading cause of neurodevelopmental impairment. Exploring the new molecular mechanism of HI brain injury has important clinical translational significance for the next clinical intervention research. Lipid phosphatase-related proteins (PLPPRs) are regulators of mitochondrial membrane integrity and energy metabolism. We recently found that Plppr5 knockout exacerbated HI impairment in some aspects and partially attenuated the neuroprotective effects of melatonin, suggesting that Plppr5 may be a novel intervention target for HI. The present study aimed to determine the long-term effects of gene knockout of Plppr5 on HI brain injury, focusing on the neuronal excitability phenotype, and to determine the effect of Plppr5 gene silencing on neuronal zinc metabolism and mitochondrial function in vitro. 10-day-old wild type (WT) mice and Plppr5-deficient (Plppr5–/–) mice were subjected to hypoxia-ischemia. Lesion volumes and HI-induced neuroexcitotoxic phenotypes were quantified together with ZnT1 protein expression in hippocampus. In addition, HT22 (mouse hippocampal neuronal cells) cell model was established by oxygen–glucose deprivation/reoxygenation (OGD/R) treatment and was treated with medium containing LV-sh_Plppr5 or control virus. Mitochondrial oxidative stress indicator ROS, mitochondrial ZnT1 protein expression and zinc ion content were detected.ResultsPlppr5-deficient mice subjected to hypoxia-ischemia at postnatal day 10 present significantly higher cerebral infarction. Plppr5-deficient mice were endowed with a more pronounced superexcitability phenotype at 4 weeks after HI, manifested as a reduced seizure threshold. ZnT1 protein was also found reduced in Plppr5-deficient mice as well as in mice subjected to HI excitotoxicity. Plppr5 knockout in vivo exacerbates HI brain injury phenotypes, including infarct volume and seizure threshold. In addition, knockout of the Plppr5 gene reduced the MFS score to some extent. In vitro Plppr5 silencing directly interferes with neuronal zinc metabolism homeostasis and exacerbates hypoxia-induced mitochondrial oxidative stress damage. Taken together, our findings demonstrate for the first time that Plppr5-deficient mouse pups exposed to neuronal hypoxia and ischemia exhibit aggravated acute brain injury and long-term brain excitability compared with the same treated WT pups, which may be related to the disruption of zinc and mitochondria-dependent metabolic pathways in the hippocampus. These data support further investigation into novel approaches targeting Plppr5-mediated zinc and mitochondrial homeostasis in neonatal HIE.

Background: Mitochondrial deficiency is a known pathology in sepsis-induced organ failure. We previously found that mitochondria-associated membranes (MAMs), a subcellular domain supporting mitochondrial status, are impaired in the heart during endotoxemia, suggesting a mechanism of mitochondrial damage occurred in sepsis. Mitophagy pathway via E3 ubiquitin ligase Parkin and PTEN-induced kinase 1 (PINK1) controls mitochondrial quality. Studies described here examined the impact of Parkin on cardiac MAMs and endotoxemia-induced cardiomyopathy. Additionally, point mutation W403A in Parkin was previously identified as a constitutively active mutation in vitro. In vivo effects of forced expression of this mutation were evaluated in the endotoxemia model.Methods: Mice of wild type (WT), Parkin-deficiency (Park2−/−), and knock-in expression of Parkin W402A (human Parkin W403A) were given lipopolysaccharide (LPS) challenge. Cardiac function was evaluated by echocardiography. In the harvested heart tissue, MAM fractions were isolated by ultracentrifugation, and their amount and function were quantified. Ultrastructure of MAMs and mitochondria was examined by electron microscopy. Mitochondrial respiratory activities were measured by enzyme assays. Myocardial inflammation was estimated by levels of pro-inflammatory cytokine IL-6. Myocardial mitophagy was assessed by levels of mitophagy factors associated with mitochondria and degrees of mitochondria-lysosome co-localization. Parkin activation, signified by phosphorylation on serine 65 of Parkin, was also evaluated.Results: Compared with WT, Park2−/− mice showed more severely impaired cardiac MAMs during endotoxemia, characterized by disrupted structure, reduced quantity, and weakened transporting function. Endotoxemia-induced cardiomyopathy was intensified in Park2−/− mice, shown by worsened cardiac contractility and higher production of IL-6. Mitochondria from the Park2−/− hearts were more deteriorated, indicated by losses in both structural integrity and respiration function. Unexpectedly, mice carrying Parkin W402A showed similar levels of cardiomyopathy and mitochondrial damage when compared with their WT counterparts. Further, Parkin W402A mutation neither enhanced mitophagy nor increased Parkin activation in myocardium under the challenge of endotoxemia.Conclusion: our results suggest that Parkin/PINK1 mitophagy participates in the regulation of cardiac MAMs during endotoxemia. Point mutation W402A (human W403A) in Parkin is not sufficient to alleviate cardiomyopathy induced by endotoxemia in vivo.

Single-cell proteins are attracting growing attention as viable alternatives for fishmeal (FM) in aquatic feed. Methanotroph (Methylococcus capsulatus, Bath) bacteria meal FeedKind® (FK) is a type of single cell protein with high protein content (75.14%) and desirable amino acids profile, produced by Methylococcus capsulatus (Bath) living on methane consumption. The present study evaluated the potential of replacing FM with FK in the diet of black sea bream (Acanthopagrus schlegelii). Five iso-energetic and iso-nitrogenous diets were designed with FK replacing 0, 4.13, 8.27, 16.53, and 24.80% FM protein in the basal diet (40% FM content), respectively. All the diets were fed to three replicates of fish (initial weight 6.56 ± 0.02 g) for 70 days. After the feeding trial, replacing dietary 8.27% FM protein with FK significantly improved the weight gain and specific growth rate of fish (P < 0.05), while other groups showed no significant difference in the growth performance (P > 0.05). The fish fed diets with 8.27 and 16.53% replacement levels exhibited significantly increased feeding rates. The 8.27% FK diet significantly increased the whole-body and muscle crude protein contents, apparent digestibility of crude lipid, foregut, and midgut amylase activities. The microvillus density in the midgut of fish fed the 24.80% FK diet significantly increased. The diet with 8.27% FK increased the serum triglyceride content of the fish, while the 24.80% FK diet reduced the serum triglyceride, total cholesterol, and low-density lipoprotein cholesterol contents of the fish. In conclusion, the results indicated that replacing dietary FM protein with up to 24.80% FK had no adverse effects on the growth of black sea bream, whilst replacing 8.27% FM protein with FK enhanced its growth performance and feed utilization.

IntroductionEpileptogenesis is understood as the plastic process that produces a persistent reorganization of the brain’s neural network after a precipitating injury (recurrent neonatal seizures, for instance) with a latent period, finally leading to neuronal hyperexcitability. Plasticity-related genes (PRGs), also known as lipid phosphate phosphatase-related proteins (PLPPRs), are regulators of mitochondrial membrane integrity and energy metabolism. This study was undertaken to determine whether PRG5 gene knockout contributes to the delayed hypersensitivity induced by developmental seizures and the aberrant sprouting of hippocampal mossy fibers, and to determine whether it is achieved through the mitochondrial pathway. Here, we developed a “twist” seizure model by coupling pilocarpine-induced juvenile seizures with later exposure to penicillin to test the long-term effects of PRG5 knockout on seizure latency through comparison with wild-type (WT) mice. Hippocampal mossy fiber sprouting (MFS) was detected by Timm staining. In order to clarify the mechanism of the adverse reactions triggered by PRG5 knockout, hippocampal HT22 neuronal cultures were exposed to glutamate, with or without PRG5 interference. Mitochondrial function, oxidative stress indicators and zinc ion content were detected.ResultsPRG5 gene knockout significantly reduced the seizure latency, and aggravated the lowered seizure threshold induced by developmental seizures. Besides, knockout of the PRG5 gene reduced the MFS scores to a certain extent. Furthermore, PRG5 gene silencing significantly increases the zinc ion content in hippocampal neurons, impairs neuronal activity and mitochondrial function, and exacerbates glutamate-induced oxidative stress damage.ConclusionIn summary, PRG5 KO is associated with significantly greater hypersusceptibility to juvenile seizures in PRG5(–/–) mice compared with WT mice. These effects may be related to the hippocampal zinc signaling. The effects do not appear to be related to changes in MFS because KO mice with juvenile seizures had the shortest seizure latencies but exhibited less MFS than WT mice with juvenile seizures.

ObjectiveWe previously demonstrated that promoting Beclin-1–dependent autophagy is cardiac protective during endotoxemia shock, suggesting that autophagy-based approaches may become a promising therapeutic strategy for sepsis. In this study, we applied both genetic and pharmacological approaches to evaluate whether Beclin-1 activation improves sepsis outcomes in a model of pneumonia-induced sepsis.MethodsSepsis was induced in mice by Klebsiella pneumoniae infection via intubation, and outcomes of clinical sickness scores, systemic infection, inflammation, survival, and pulmonary pathology were examined. Evaluation of Beclin-1 activation was achieved by comparing strains of C57BL/6J wild type and Becn1F121A that carries a transgenic expression of Beclin-1–active mutant F121A, and by comparing animal groups treated with Beclin-1–activating peptide, Tat-beclin-1 peptide (TB-peptide), or with vehicle control. The status of autophagy in the lung tissue was examined in autophagy reporter mice, CAG-RFP-EGFP-LC3, by fluorescence microscopy.ResultsPulmonary infection by K. pneumoniae produced an insufficient, maladaptive autophagy in the lung. Activation of Beclin-1 by forced expression of active mutant Becn1F121A or by treatment with TB-peptide enhanced autophagy and significantly reduced sickness scores, systemic infection, and circulating and pulmonary cytokine production. Both approaches demonstrated notable benefits in limiting post-infection pathogenesis in the lung, such as decreases in alveolar congestion, hemorrhage, infiltration of inflammatory cells, and alveolar wall thickness.ConclusionData suggest that targeted activation of Beclin-1 alleviates adverse outcomes of pneumonia-induced sepsis, and thus, possess a therapeutic potential.

Cerebral palsy (CP), a group of clinical syndromes caused by non-progressive brain damage in the developing fetus or infant, is one of the most common causes of lifelong physical disability in children in most countries. At present, many researchers believe that perinatal cerebral hypoxic ischemic injury or inflammatory injury are the main causes of cerebral palsy. Previous studies including our works confirmed that melatonin has a protective effect against convulsive brain damage during development and that it affects the expression of various molecules involved in processes such as metabolism, plasticity and signaling in the brain. Integral membrane protein plppr5 is a new member of the plasticity-related protein family, which is specifically expressed in brain and spinal cord, and induces filopodia formation as well as neurite growth. It is highly expressed in the brain, especially in areas of high plasticity, such as the hippocampus. The signals are slightly lower in the cortex, the cerebellum, and in striatum. Noteworthy, during development plppr5 mRNA is expressed in the spinal cord, i.e., in neuron rich regions such as in medial motor nuclei, suggesting that plppr5 plays an important role in the regulation of neurons. However, the existing literature only states that plppr5 is involved in the occurrence and stability of dendritic spines, and research on its possible involvement in neonatal ischemic hypoxic encephalopathy has not been previously reported. We used plppr5 knockout (plppr5−/−) mice and their wild-type littermates to establish a model of hypoxicischemic brain injury (HI) to further explore the effects of melatonin on brain injury and the role of plppr5 in this treatment in an HI model, which mainly focuses on cognition, exercise, learning, and memory. All the tests were performed at 3–4 weeks after HI. As for melatonin treatment, which was performed 5 min after HI injury and followed by every 24h. In these experiments, we found that there was a significant interaction between genotype and treatment in novel object recognition tests, surface righting reflex tests and forelimb suspension reflex tests, which represent learning and memory, motor function and coordination, and the forelimb grip of the mice, respectively. However, a significant main effect of genotype and treatment on performance in all behavioral tests were observed. Specifically, wild-type mice with HI injury performed better than plppr5−/− mice, regardless of treatment with melatonin or vehicle. Moreover, treatment with melatonin could improve behavior in the tests for wild-type mice with HI injury, but not for plppr5−/− mice. This study showed that plppr5 knockout aggravated HI damage and partially weakened the neuroprotection of melatonin in some aspects (such as novel object recognition test and partial nerve reflexes), which deserves further study.

Purpose: Cells recognize mechanical cues from the extracellular environment through interactions with matrix proteins, such as collagen. Following injury, fibrotic deposition of collagen can lead to aberrant cell differentiation and failed healing. Aligned collagen matrices can drive PDGFRα+ mesenchymal progenitor cells (MPCs) towards osteochondral lineages, leading to heterotopic ossification (HO). Previous work has shown that reducing mechanical strain by immobilizing a limb prevents the formation of an aligned extracellular matrix (ECM), leading to altered MPC differentiation, however, the cellular mechanisms of matrix reorganization following injury remain unclear. Using novel bioinformatics approach, we identified discoidin domain receptor 2 (DDR2) as a key MPC specific tyrosine kinase receptor that interacts with the fibrillar collagen matrix. We hypothesized that DDR2 activation leads to regulation of ECM alignment and therefore serves as a novel upstream regulator of mechanotransductive signaling following musculoskeletal repair. Methods: Heterotopic ossification was induced using a proven mouse model of 30% total body surface area burn with concurrent Achilles’ transection in Ddr2 deletion (Ddr2slie/slie) and littermate control mice on the C57/BL6J background. In separate experiments, ankle joint immobilizers were placed on injured mice for 1, 2 or 3 weeks after burn/tenotomy. Tissue from Ddr2 deleted, immobilized and mobile control mice were harvested for immunofluorescence histology, second harmonic generation of collagen fiber histology at 1, 3 and 9 weeks after injury (SHG) (n=4/group). HO site tissue was also harvested prior to injury and at 1 and 6 weeks after burn/tenotomy and processed for Single-Cell RNA sequencing (scRNA seq) and single nucleus assay for transposase-accessible chromatin (snATAC) using 10X Sequencing and downstream analysis using Seurat 4.0.1 and Signac 3.1.5. Results: Single-cell RNA sequencing showed that genes for collagen type 1 and 3 and Ddr2 are highly upregulated in MPCs following injury. SnATAC showed increased open chromatin reads in the promoter of MPCs following injury (not shown). Utilizing SHG, we found that the collagen matrix is aligned at three-weeks following injury, but one week of limb immobilization was enough to prevent aligned collagen. The active phosphorylated form of DDR2 (pDDR2) was decreased in PDGFRα+ MPCs within immobilized samples, suggesting that DDR2 may affect matrix alignment. Ddr2slie/slie had more disorganized collagen matrix nine weeks following injury and significantly reduced HO. Conclusions: This is the first work to alter collagen matrix organization by genetic modification and mobilization protocols and identifies DDR2 as a critical upstream receptor in matrix organization and aberrant progenitor differentiation following injury. DDR2 antagonism could have therapeutic potential in preventing heterotopic ossification following traumatic injury.

Mesenchymal stem cells (MSCs) derived from different tissues may aid in the regeneration of radiation-induced organ lesions; however, the radiation responses of human MSCs from different sources are unknown. In our study, a comparison of the results from cell proliferation, apoptosis, cell cycle, DNA damage, and DNA repair assays consistently showed that MSCs derived from adipose tissue possess a significantly stronger radiation resistance capacity than MSCs derived from umbilical cord and gingival, which is accompanied by a higher level of phosphorylated signal transducer and activator of transcription 3 (Stat3) expression. This reminds us Stat3 could be a potential biomarker of radiation resistance. These findings provide a better understanding of radiation-induced biologic responses in MSCs and may lead to the development of better strategies for stem cell treatment and cancer therapy.

Abstract Background Behavioral assessment has been acted as the gold standard for the diagnosis of disorders of consciousness (DOC) patients. The item “Functional Object Use” in the motor function sub-scale in the Coma Recovery Scale-Revised (CRS-R) is a key item in differentiating between minimally conscious state (MCS) and emergence from MCS (EMCS). However, previous studies suggested that certain specific stimuli, especially something self-relevant can affect DOC patients’ scores of behavioral assessment scale. So, we attempted to find out if personalized objects can improve the diagnosis of EMCS in the assessment of Functional Object Use by comparing the use of patients’ favorite objects and other common objects in MCS patients. Methods Twenty-one post-comatose patients diagnosed as MCS were prospectively included. The item “Functional Object Use” was assessed by using personalized objects (e.g., cigarette, paper) and non-personalized objects, which were presented in a random order. The rest assessments were performed following the standard protocol of the CRS-R. The differences between functional uses of the two types of objects were analyzed by the McNemar test. Results The incidence of Functional Object Use was significantly higher using personalized objects than non-personalized objects in the CRS-R. Five out of the 21 MCS studied patients, who were assessed with non-personalized objects, were re-diagnosed as EMCS with personalized objects (χ2 = 5, df = 1, p

Acidification (OA), a global threat to the world’s oceans, is projected to significantly grow if CO2 continues to be emitted into the atmosphere at high levels. This will result in a slight decrease in pH. Since the latter is a logarithmic scale of acidity, the higher acidic seawater is expected to have a tremendous impact on marine living resources in the long-term. An 8-week laboratory experiment was designed to assess the impact of the projected pH in 2100 and beyond on fish survival, health, growth, and fish meat quality. Two projected scenarios were simulated with the control treatment, in triplicates. The control treatment had a pH of 8.10, corresponding to a pCO2 of 321.37 ± 11.48 µatm. The two projected scenarios, named Predict_A and Predict_B, had pH values of 7.80-pCO2 = 749.12 ± 27.03 and 7.40-pCO2 = 321.37 ± 11.48 µatm, respectively. The experiment was preceded by 2 weeks of acclimation. After the acclimation, 20 juvenile black sea breams (Acanthopagrus schlegelii) of 2.72 ± 0.01 g were used per tank. This species has been selected mainly due to its very high resistance to diseases and environmental changes, assuming that a weaker fish resistance will also be susceptibly affected. In all tanks, the fish were fed with the same commercial diet. The seawater’s physicochemical parameters were measured daily. Fish samples were subjected to physiological, histological, and biochemical analyses. Fish growth, feeding efficiency, protein efficiency ratio, and crude protein content were significantly decreased with a lower pH. Scanning electron microscopy revealed multiple atrophies of microvilli throughout the small intestine’s brush border in samples from Predict_A and Predict_B. This significantly reduced nutrient absorption, resulting in significantly lower feed efficiency, lower fish growth, and lower meat quality. As a result of an elevated pCO2 in seawater, the fish eat more than normal but grow less than normal. Liver observation showed blood congestion, hemorrhage, necrosis, vacuolation of hepatocytes, and an increased number of Kupffer cells, which characterize liver damage. Transmission electron microscopy revealed an elongated and angular shape of the mitochondrion in the liver cell, with an abundance of peroxisomes, symptomatic of metabolic acidosis.

Trichomes are specialized structures that originate from epidermal cells of organs in higher plants. The cotton fiber is a unique single-celled trichome that elongates from the seed coat epidermis. Cotton (Gossypium hirsutum) fibers and trichomes are models for cell differentiation. In an attempt to elucidate the intercellular factors that regulate fiber and trichome cell development, we identified a plasmodesmal b-1,3-glucanase gene (designated GhPdBG) controlling the opening and closing of plasmodesmata in cotton fibers. Structural and evolutionary analysis showed haplotypic variation in the promoter region of the GhPdBG gene among 352 cotton accessions, but high conservation in the coding region. GhPdBG was expressed predominantly in cotton fibers and localized to plasmodesmata (PD). Expression patterns of PdBG that corresponded to PD permeability were apparent during fiber development in G. hirsutum and G. barbadense. The PdBG-mediated opening-closure of PD appears to be involved in fiber development and may account for the contrasting fiber traits of these two species. Ectopic expression of GhPdBG revealed that it functions in regulating fiber and trichome length and/or density by modulating plasmodesmatal permeability. This finding suggests that plasmodesmal targeting of GhPdBG, as a switch of intercellular channels, regulates single-celled fiber and trichome development in cotton. [ABSTRACT FROM AUTHOR]

Efforts to identify and target glioblastoma (GBM) drivers have primarily focused on receptor tyrosine kinases (RTKs). Clinical benefits, however, have been elusive. Here, we identify an SRY-related box 2 (SOX2) transcriptional regulatory network that is independent of upstream RTKs and capable of driving glioma-initiating cells. We identified oligodendrocyte lineage transcription factor 2 (OLIG2) and zinc-finger E-box binding homeobox 1 (ZEB1), which are frequently co-expressed irrespective of driver mutations, as potential SOX2 targets. In murine glioma models, we show that different combinations of tumor suppressor and oncogene mutations can activate Sox2, Olig2, and Zeb1 expression. We demonstrate that ectopic co-expression of the three transcription factors can transform tumor-suppressor-deficient astrocytes into glioma-initiating cells in the absence of an upstream RTK oncogene. Finally, we demonstrate that the transcriptional inhibitor mithramycin downregulates SOX2 and its target genes, resulting in markedly reduced proliferation of GBM cells in vivo.

Objective: To detect the initiation of swallowing in patients with disorders of consciousness (DOC) as well as the relationship between the initiation of swallowing and the prognosis of DOC patients.Methods: Nineteen DOC patients were included in this study, and a self-controlled trial compared five different stimuli. The five different stimuli were as follows: (1) one command, as recommended by the Coma Recovery Scale-Revised (CRS-R), which was “open your mouth”; (2) placing a spoon in front of the patient's mouth without a command; (3) placing a spoon filled with water in front of the patient's mouth without a command; (4) one command—“there is a spoon; open your mouth”—with a spoon in front of the patient's mouth; (5) one command, “there is a spoon with water; open your mouth,” with a spoon filled with water in front of the patient's mouth. All 19 patients were given these five stimuli randomly, and any one of the commands was presented four times to a patient, one at a time, at 15-s intervals. The sensitivity and specificity of the initiation of swallowing in detecting conscious awareness were determined.Results: None of the patients responded to the first four stimuli. However, six patients showed initiated swallowing toward the fifth stimulus. Among those six, five patients showed improvement in their consciousness state 6 months later. The sensitivity and specificity of the initiation of swallowing for DOC patients was 83.33% [95% CIs (36%, 100%)] and 92.31% [95% CIs (64%, 100%)], respectively.Conclusions: The initiation of swallowing can be an early indication of conscious behavior and can likely provide evidence of conscious awareness.Clinical Trial Registration:www.ClinicalTrials.gov, identifier: NCT03508336; Date of registration: 2018/4/16.

Radiation therapy is one of the most common cancer treatments. It is important to understand how cells respond to ionizing radiation (IR) to improve therapeutic efficacy. Circular RNAs (circRNAs) recently have been found to regulate a variety of cellular processes. However, it is poorly defined that their expression pattern and their identity in cells following IR exposure. Here, we performed high-throughput sequencing and comprehensive analysis of circRNA expression in human embryonic kidney (HEK) 293T cells before and after irradiation. We identified totally 5592 circRNAs and discovered 1038 new circRNAs. We found 158 circRNAs with significantly differential expression after IR exposure. Among them, there were 61 upregulated and 97 downregulated circRNAs. Using Gene Ontology, Kyoto Encyclopedia of Genes and Genomes pathway, and circRNA-microRNA-messenger RNA network analyses, we found the differentially expressed circRNAs might be involved in the signal pathways of oxidative phosphorylation, epithelial growth factor receptor (EGFR) tyrosine kinase inhibitor resistance, and mammalian target of rapamycin (mTOR) signaling.

Sepsis is a leading cause of death in intensive care units, and cardiac dysfunction is an identified serious component of the multi-organ failure associated with this critical condition. This review summarized the current discoveries and hypotheses of how autophagy changes in the heart during sepsis and the underlying mechanisms. Recent investigations suggest that specific activation of autophagy initiation factor Beclin-1 has a potential to protect cardiac mitochondria, attenuate inflammation, and improve cardiac function in sepsis. Accordingly, pharmacological interventions targeting this pathway have a potential to become an effective approach to control sepsis outcomes. The role of autophagy during sepsis pathogenesis has been under intensive investigation in recent years. It is expected that developing therapeutic approaches with specificities targeting at autophagy regulatory factors may provide new opportunities to alleviate organ dysfunction caused by maladaptive autophagy during sepsis.

We have previously shown that mitochondria-targeted vitamin E (Mito-Vit-E), a mtROS specific antioxidant, improves cardiac performance and attenuates inflammation in a pneumonia-related sepsis model. In this study, we applied the same approaches to decipher the signaling pathway(s) of mtROS-dependent cardiac inflammation after sepsis. Sepsis was induced in Sprague Dawley rats by intratracheal injection of S. pneumoniae. Mito-Vit-E, vitamin E or vehicle was administered 30 minutes later. In myocardium 24 hours post-inoculation, Mito-Vit-E, but not vitamin E, significantly protected mtDNA integrity and decreased mtDNA damage. Mito-Vit-E alleviated sepsis-induced reduction in mitochondria-localized DNA repair enzymes including DNA polymerase γ, AP endonuclease, 8-oxoguanine glycosylase, and uracil-DNA glycosylase. Mito-Vit-E dramatically improved metabolism and membrane integrity in mitochondria, suppressed leakage of mtDNA into the cytoplasm, inhibited up-regulation of Toll-like receptor 9 (TLR9) pathway factors MYD88 and RAGE, and limited RAGE interaction with its ligand TFAM in septic hearts. Mito-Vit-E also deactivated NF-κB and caspase 1, reduced expression of the essential inflammasome component ASC, and decreased inflammatory cytokine IL-1β. In vitro, both Mito-Vit-E and TLR9 inhibitor OND-I suppressed LPS-induced up-regulation in MYD88, RAGE, ASC, active caspase 1, and IL-1β in cardiomyocytes. Since free mtDNA escaped from damaged mitochondria function as a type of DAMPs to stimulate inflammation through TLR9, these data together suggest that sepsis-induced cardiac inflammation is mediated, at least partially, through mtDNA-TLR9-RAGE. At last, Mito-Vit-E reduced the circulation of myocardial injury marker troponin-I, diminished apoptosis and amended morphology in septic hearts, suggesting that mitochondria-targeted antioxidants are a potential cardioprotective approach for sepsis.