Paediatric sepsis-associated encephalopathy (SAE): a comprehensive review.

Sepsis-associated encephalopathy (SAE) is one of the most common types of organ dysfunction without overt central nervous system (CNS) infection. It is associated with higher mortality, low quality of life, and long-term neurological sequelae, its mortality in patients diagnosed with sepsis, progressing to SAE, is 9% to 76%. The pathophysiology of SAE is still unknown, but its mechanisms are well elaborated, including oxidative stress, increased cytokines and proinflammatory factors levels, disturbances in the cerebral circulation, changes in blood-brain barrier permeability, injury to the brain's vascular endothelium, altered levels of neurotransmitters, changes in amino acid levels, dysfunction of cerebral microvascular cells, mitochondria dysfunction, activation of microglia and astrocytes, and neuronal death. The diagnosis of SAE involves excluding direct CNS infection or other types of encephalopathies, which might hinder its early detection and appropriate implementation of management protocols, especially in paediatric patients where only a few cases have been reported in the literature. The most commonly applied diagnostic tools include electroencephalography, neurological imaging, and biomarker detection. SAE treatment mainly focuses on managing underlying conditions and using antibiotics and supportive therapy. In contrast, sedative medication is used judiciously to treat those showing features such as agitation. The most widely used medication is dexmedetomidine which is neuroprotective by inhibiting neuronal apoptosis and reducing a sepsis-associated inflammatory response, resulting in improved short-term mortality and shorter time on a ventilator. Other agents, such as dexamethasone, melatonin, and magnesium, are also being explored in vivo and ex vivo with encouraging results. Managing modifiable factors associated with SAE is crucial in improving generalised neurological outcomes. From those mentioned above, there are still only a few experimentation models of paediatric SAE and its treatment strategies. Extrapolation of adult SAE models is challenging because of the evolving brain and technical complexity of the model being investigated. Here, we reviewed the current understanding of paediatric SAE, its pathophysiological mechanisms, diagnostic methods, therapeutic interventions, and potential emerging neuroprotective agents.

Melatonin Promotes Mitochondrial Biogenesis and Mitochondrial Degradation in Hepatocytes During Sepsis.

Objective: This study aimed to investigate the protective mechanisms of melatonin in an in vitro model of sepsis-induced hepatocyte injury, specifically focusing on mitophagy and mitochondrial biogenesis. Methods: In this study, we utilized lipopolysaccharide (LPS)-treated AML12 cells to establish an in vitro model of sepsis-induced hepatocyte injury. The effects of melatonin pretreatment were examined through various analyses, including assessments of oxidative stress, inflammation, mitophagy, mitochondrial biogenesis, and adenosine triphosphate (ATP) levels. Results: The results revealed that LPS-treated AML12 cells exhibited elevated levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6 protein, intracellular reactive oxygen species (ROS), and lipid peroxidation, specifically malondialdehyde (MDA). Moreover, the levels of key markers associated with mitophagy, including PTEN-induced putative kinase 1 (PINK1), parkin, and LC3, were significantly increased (P < .05). Similarly, markers of mitochondrial biogenesis, such as peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α), nuclear respiratory factor 1 (NRF1), and mitochondrial transcription factor A (TFAM), were also significantly increased (P < .05). Conversely, superoxide dismutase (SOD) activity and ATP levels were significantly decreased in LPS-treated AML12 cells compared to the control group (P < .05). However, melatonin pretreatment led to a significant decrease in TNF-α and IL-6 protein levels, intracellular ROS, and MDA levels (P < .05), along with a significant increase in SOD activity, ATP levels, and markers of mitophagy and mitochondrial. Conclusions: Our findings demonstrate that melatonin plays a role in regulating mitochondrial quality control in sepsis-induced hepatocytes. It achieves this result by promoting mitophagy and inducing mitochondrial biogenesis, thereby selectively eliminating dysfunctional mitochondria.

Effects of hydrogen-rich saline in neuroinflammation and mitochondrial dysfunction in rat model of sepsis-associated encephalopathy.

BACKGROUND: Sepsis-associated encephalopathy (SAE) is one of the most common types of sepsis-related organ dysfunction without overt central nervous system (CNS) infection. It is associated with higher mortality, low quality of life, and long-term neurological sequelae in suspected patients. At present there is no specific treatment for SAE rather than supportive therapy and judicious use of antibiotics, which are sometimes associated with adverse effects. Molecular hydrogen (H2) has been reported to play crucial role in regulating inflammatory responses, neuronal injury, apoptosis and mitochondrial dysfunction in adult models of SAE. Here we report the protective effect of hydrogen-rich saline in juvenile SAE rat model and its possible underling mechanism(s). MATERIALS AND METHODS: Rats were challenged with lipopolysaccharide (LPS) at a dose of 8 mg/kg injected intraperitoneally to induce sepsis and hydrogen-rich saline (HRS) administered 1 h following LPS induction at a dose of 5 ml/kg. Rats were divided into: sham, sham + HRS, LPS and LPS + HRS. At 48 h, rats were sacrificed and Nissl staining for neuronal injury, TUNEL assay for apoptotic cells detection, immunohistochemistry, and ELISA protocol for inflammatory cytokines determination, mitochondrial dysfunction parameters, electron microscopy and western blot analysis were studied to examine the effect of HRS in LPS-induced septic rats. RESULTS: Rats treated with HRS improved neuronal injury, improvement in rats' survival rate. ELISA analysis showed decreased TNF-α and IL-1ß and increased IL-10 expression levels in the HRS-treated group. Apoptotic cells were decreased after HRS administration in septic rats. The numbers of GFAP and IBA-1positive cells were attenuated in the HRS-treated group when compared to the LPS group. Subsequently, GFAP and IBA-1 immunoreactivity were decreased after HRS treatment. Mitochondrial membrane potential detected by JC-1 dye and ATP content were decreased in septic rats, which were improved after HRS treatment, while release of ROS was increased in the LPS group reverted by HRS treatment, ameliorating mitochondrial dysfunction. Further analysis by transmission electron microscopy showed decreased number of mitochondria and synapses, and disrupted mitochondrial membrane ultrastructure in the LPS group, while HRS administration increased mitochondria and synapses number. CONCLUSION: These data demonstrated that HRS can improve survival rate, attenuate neuroinflammation, astrocyte and microglial activation, neuronal injury and mitochondrial dysfunction in juvenile SAE rat model, making it a potential therapeutic candidate in treating paediatric SAE.

Persistent viral shedding of human adenovirus type 7 in children with severe pneumonia.

To understand host-pathogen interactions and develop effective prevention and control strategies for human adenovirus (HAdV), it is essential to explore the characteristics of HAdV shedding. Hospitalized children <14 years who had severe HAdV pneumonia were tested for HAdV DNA by quantitative real-time PCR in nasopharyngeal aspirate (NPA). A total of 132 children were enrolled, including 102 patients with HAdV type 7 (HAdV-7) infection and 12 patients with HAdV type 3 (HAdV-3) infection. A total of 1372 qualified NPA samples were collected. There was a significant negative correlation between the viral load of HAdV and the course of the disease (Spearman r = -0.547, p = .000). HAdV-7 load decreased at a rate of 0.089 log10 copies/mL per day (95% CI: -0.096 to -0.081; R 2 = 0.332), and the duration of viral shedding was predicted to be 96.9 days (y = 8.624-0.089x). However, HAdV-3 load decreased more quickly (95% CI: - 0.229 to - 0.143; R 2 = 0.403), and the duration of viral shedding was 51.4 days (y = 9.558-0.186x). The median viral load of the HAdV-7 group at weeks 2 and 3, and more than 3 weeks postinfection was higher than that of the HAdV-3 group. No significant differences in the duration of viral shedding were found in different gender, age (>2 vs. ≤2 years), and with or without underlying diseases groups. Viral shedding in children with severe HAdV pneumonia persisted, among which HAdV-7 lasted longer than 3 months and the viral load decreased slowly than HAdV-3.

Continuous Renal-Replacement Therapy in Critically Ill Children: Practice Changes and Association With Outcome.

OBJECTIVES: This study was designed to evaluate practice changes and outcomes over a 10-year period in a large single-center PICU cohort that received continuous renal-replacement therapy. DESIGN: Retrospective study design. SETTING: A multidisciplinary tertiary PICU of a university-affiliated hospital in Guangzhou, China. PATIENTS: All critically ill children who were admitted to our PICU from January 2010 to December 2019 and received continuous renal-replacement therapy were included in this study. MEASUREMENTS AND MAIN RESULTS: A total of 289 patients were included in the study. Of the two study periods, 2010-2014 and 2015-2019, the proportion of continuous renal-replacement therapy initiation time greater than 24 hours was significantly reduced ([73/223] 32.73% vs. [40/66] 60.60%, p < 0.001), the percentage of fluid overload at continuous renal-replacement therapy initiation was lower (3.8% [1.6-7.2%] vs. 12.1% [6.6-23.3%], p < 0.001), the percentage of regional citrate anticoagulation protocol was increased ([223/223] 100% vs. [15/66] 22.7%, p < 0.001), and the ICU survival rate was significantly improved ([24/66] 36.4% vs. [131/223] 58.7%, p = 0.001) in the latter period compared with the former. In addition, subgroup analysis found that survival were higher in patients with continuous renal-replacement therapy initiation time less than 24 hours, regional citrate anticoagulation protocol, and fluid overload less than 10%. CONCLUSIONS: The survival rate of patients received continuous renal-replacement therapy treatment in our center has improved over past 10 years, and some changes have taken place during these periods. Among them, early initiation of continuous renal-replacement therapy, lower fluid overload, and regional citrate anticoagulation method seems to be related to the improvement of outcome. Ongoing evaluation of the practice changes and quality improvement of continuous renal-replacement therapy for critically ill pediatric patients still need attention.

Genome-wide CRISPR-Cas9 viability screen reveals genes involved in TNF-α-induced apoptosis of human umbilical vein endothelial cells.

Tumor necrosis factor α (TNF-α), a pivotal cytokine in sepsis, protects the host against pathogens by promoting an inflammatory response while simultaneously inducing apoptosis of the vascular endothelium. Unfortunately, inhibitors targeting certain components of the TNF-α signaling pathway to reduce cellular apoptosis have failed to translate into clinical applications, partly due to the adverse effects of excessive immunosuppression. In an attempt to discover potential targets in the TNF-α signaling pathway to modulate moderate inflammation and apoptosis during the development of sepsis, we performed a pooled genome-wide CRISPR/Cas9 knockout screen in human umbilical vein endothelial cells (HUVECs). Tumor necrosis factor receptor superfamily member 1A (TNFRSF1A), B-cell lymphoma 2 (BCL2), Bcl2-associated death promoter (BAD), and NLR family member X1 (NLRX1) deficiencies were identified as the effective genetic suppressors of TNF-α cytotoxicity on a list of candidate regulators. CRISPR-mediated NLRX1 knockout conferred cellular resistance to challenge with TNF-α, and NLRX1 could be induced to colocalize with mitochondria following TNF-α stimulation. Thus, our work demonstrates the advantage of genome-scale screening with Cas9 and validates NLRX1 as a potential modulator of TNF-α-induced vascular endothelial apoptosis during sepsis.

Tumor-targeted in vivo gene silencing via systemic delivery of cRGD-conjugated siRNA.

RNAi technology is taking strong position among the key therapeutic modalities, with dozens of siRNA-based programs entering and successfully progressing through clinical stages of drug development. To further explore potentials of RNAi technology as therapeutics, we engineered and tested VEGFR2 siRNA molecules specifically targeted to tumors through covalently conjugated cyclo(Arg-Gly-Asp-d-Phe-Lys[PEG-MAL]) (cRGD) peptide, known to bind αvß3 integrin receptors. cRGD-siRNAs were demonstrated to specifically enter and silence targeted genes in cultured αvß3 positive human cells (HUVEC). Microinjection of zebrafish blastocysts with VEGFR2 cRGD-siRNA resulted in specific inhibition of blood vessel growth. In tumor-bearing mice, intravenously injected cRGD-siRNA molecules generated no innate immune response and bio-distributed to tumor tissues. Continuous systemic delivery of two different VEGFR2 cRGD-siRNAs resulted in down-regulation of corresponding mRNA (55 and 45%) and protein (65 and 45%) in tumors, as well as in overall reduction of tumor volume (90 and 70%). These findings demonstrate strong potential of cRGD-siRNA molecules as anti-tumor therapy.

Using a practical molecular capsular serotype prediction strategy to investigate Streptococcus pneumoniae serotype distribution and antimicrobial resistance in Chinese local hospitalized children.

BACKGROUND: China is one of ten countries with the highest prevalence rate of pneumococcal infections. However, there is limited serotype surveillance data for Streptococcus pneumoniae, especially from the community or rural regions, partly due to limited serotyping capacity because Quellung serotyping is only available in few centers in China. The aim of this study was to develop a simple, practical and economic pneumococcal serotype prediction strategy suitable for future serotype surveillance in China. METHODS: In this study, 193 S. pneumoniae isolates were collected from hospitalized children, 96.9 % of whom were < 5 years old. The cpsB sequetyping, complemented by selective and modified USA CDC sequential multiplex-PCR, was performed on all the isolates, and serotypes 6A-6D specific PCRs were done on all serogroup 6 isolates. Based on systematic analysis of available GenBank cpsB sequences, we established a more comprehensive cpsB sequence database than originally published for cpsB sequetyping. Antibiotic susceptibility of all isolates was determined using the disk diffusion or E-test assays. RESULTS: We built up a comprehensive S. pneumoniae serotype cpsB sequetyping database for all the 95 described serotypes first, and then developed a simple strategy for serotype prediction based on the improved cpsB sequetyping and selective multiplex-PCR. Using the developed serotype prediction strategy, 191 of 193 isolates were successfully "serotyped", and only two isolates were "non-serotypeable". Sixteen serotypes were identified among the 191 "serotypeable" isolates. The serotype distribution of the isolates from high to low was: 19 F (34.7 %), 23 F (17.1 %), 19A (11.9 %), 14 (7.3 %), 15B/15C (6.7 %), 6B (6.7 %), 6A (6.2 %), 9 V/9A (1.6 %); serotypes 6C, 3, 15 F/15A, 23A and 20 (each 1.1 %); serotypes 10B, 28 F/28A and 34 (each 0.5 %). The prevalence of parenteral penicillin resistance was 1.0 % in the non-meningitis isolates and 88.6 % in meningitis isolates. The total rate of multidrug resistance was 86.8 %. CONCLUSIONS: The integrated cpsB sequetyping supplemented with selective mPCR and serotypes 6A-6D specific PCRs "cocktail" strategy is practical, simple and cost-effective for use in pneumococcal infection serotype surveillance in China. For hospitalized children with non-meningitis penicillin-susceptible pneumococcal infections, clinicians still can use narrow-spectrum and cheaper penicillin, using the parenteral route, rather than using broader-spectrum and more expensive antimicrobials.

[Correlation between uncoupling protein 2 expression and myocardial mitochondrial injury in rats with sepsis induced by lipopolysaccharide].

OBJECTIVE: To investigate the correlation between uncoupling protein 2 (UCP2) expression and myocardial mitochondria injury in rats with sepsis induced by lipopolysaccharide (LPS). METHODS: The rat model of sepsis was established through an intraperitoneal injection of LPS. Forty male Sprague-Dawley rats were randomly and equally divided into control group (an intraperitoneal injection of normal saline), sepsis 6 h group (LPS-6 h group), sepsis 12 h group (LPS-12 h group), sepsis 24 h group (LPS-24 h group), and sepsis 48 h group (LPS-48 h group). The serum and heart tissues were harvested at corresponding time points and myocardial mitochondria was extracted. The microplate reader was applied to measure creatine kinase (CK), creatine kinase-MB (CK-MB), and reactive oxygen species (ROS). Flow cytometry was applied to measure the degree of mitochondrial swelling and mitochondrial membrane potential (MMP). Western blot was used to measure the expression level of UCP2. Electron microscopy was applied to observe the morphological changes in heart tissues and myocardial mitochondria. RESULTS: Compared with the control group, the LPS groups had significantly increased serum levels of CK, CK-MB, and myocardial ROS, as well as a significantly increased degree of mitochondrial swelling (P<0.05), and these values reached their peaks at 24 hours after LPS injection. The LPS groups had a significant decrease in MMP (P<0.05), which reached the lowest level at 24 hours after LPS injection. Western blot showed that the LPS groups had a significant increase in the expression level of myocardial UCP2 compared with the control group (P<0.05), which reached its peak at 24 hours after LPS injection. The results of electron microscopy showed mitochondrial swelling, partial rupture of the mitochondrial membrane, and cavity formation in rats in the LPS groups. The most severe lesions occurred in the LPS-24 h group. In rats with LPS, the ROS level in the myocardial mitochondria and the degree of mitochondrial swelling were positively correlated with the expression level of UCP2 (r=0.796 and 0.893, respectively; P<0.05), while MMP was negatively correlated with the expression level of UCP2 (r=-0.903, P<0.05). CONCLUSIONS: In the rat model of sepsis, the myocardium and myocardial mitochondria have obvious injuries, and the expression level of UCP2 is closely correlated with mitochondrial injury. Therefore, UCP2 might play an important role in myocardial mitochondrial injury in sepsis.

Experimental treatments for mitochondrial dysfunction in sepsis: A narrative review.

Sepsis is a systemic inflammatory response to infection. Sepsis, which can lead to severe sepsis, septic shock, and multiple organ dysfunction syndrome, is an important cause of mortality. Pathogenesis is extremely complex. In recent years, cell hypoxia caused by mitochondrial dysfunction has become a hot research field. Sepsis damages the structure and function of mitochondria, conversely, mitochondrial dysfunction aggravated sepsis. The treatment of sepsis lacks effective specific drugs. The aim of this paper is to undertake a narrative review of the current experimental treatment for mitochondrial dysfunction in sepsis. The search was conducted in PubMed databases and Web of Science databases from 1950 to January 2014. A total of 1,090 references were retrieved by the search, of which 121 researches met all the inclusion criteria were included. Articles on the relationship between sepsis and mitochondria, and drugs used for mitochondrial dysfunction in sepsis were reviewed retrospectively. The drugs were divided into four categories: (1) Drug related to mitochondrial matrix and respiratory chain, (2) drugs of mitochondrial antioxidant and free radical scavengers, (3) drugs related to mitochondrial membrane stability, (4) hormone therapy for septic mitochondria. In animal experiments, many drugs show good results. However, clinical research lacks. In future studies, the urgent need is to develop promising drugs in clinical trials.

[A preliminary study of long-term mitochondrial dysfunction in rat brain caused by lipopolysaccharide-induced sepsis].

OBJECTIVE: To preliminarily investigate the long-term structural and functional injuries of mitochondria in rat brain caused by sepsis. METHODS: Wistar rats were randomly assigned into sepsis and control groups. A rat model of sepsis was prepared by an intraperitoneal injection of 10 mg/kg lipopolysaccharide (LPS) of gram-negative bacteria, and the survival assay was performed. Eight rats in the sepsis group were sacrificed at 12, 24, 48, or 72 hours after LPS injection, while rats in the control group were sacrificed after an intraperitoneal injection of an equal volume of normal saline. Mitochondria were extracted from rat brain tissue. Mitochondrial membrane potential (MMP) and mitochondrial swelling level were determined by flow cytometry, and the activities of electron transport chain complexes (I-V) were measured using enzyme assay kits. Hematoxylin-eosin (HE) staining and electron microscopy were used to observe morphological changes in brain tissue and mitochondria. RESULTS: The sepsis group had a significantly lower survival rate than the control group (P<0.01). The MMP and activities of electron transport chain complexes (I-V) in the sepsis group, which were significantly lower than those in the control group (P<0.05), were reduced to the lowest levels at 48 hours and partially recovered at 72 hours. The mitochondrial swelling level in the sepsis group, which was significantly higher than that in the control group (P<0.05), increased to the peak level at 48 hours and partially recovered at 72 hours. Hematoxylin and Eosin staining revealed substantial damages in the structure of brain tissue, and electron microscopy showed mitochondrial swelling, and vacuolization in a few mitochondria. CONCLUSIONS: In the rat model of LPS-induced sepsis, both structural and functional injuries are found in cerebral mitochondria, and achieve the peak levels probably at around 48 hours.

[Effect of UCP2-siRNA on inflammatory response of cardiomyocytes induced by septic serum].

OBJECTIVE: To study the effect of uncoupling protein 2 (UCP2)-siRNA on the inflammatory response of rat cardiomyocytes (H9C2) induced by septic serum and to investigate the possible role of UCP2 in the development of septic cardiomyopathy. METHODS: Serum samples were separately collected from normal rats and septic rats. Cultured rat cardiac cells (H9C2) were randomly divided into blank control, normal serum, 10% septic serum, UCP2-siRNA+10% septic serum and negative siRNA+10% septic serum groups. Stimulation with 10% septic serum was performed for 12 hours in relevant groups. The mRNA expression of tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) was measured by RT-PCR. The expression of phosphorylated p38 mitogen-activated protein kinase (p-p38 MAPK) and nuclear factor-kappa B (NF-κB) was measured by Western blot. RESULTS: The expression levels of p-p38 and NF-κB in the UCP2-siRNA+10% septic serum group were significantly higher than in the 10% septic serum group (P<0.05). The UCP2-siRNA+10% septic serum group had a significantly higher TNF-α mRNA expression than the 10% septic serum group (P<0.01), but IL-1ß mRNA expression showed no significant difference between the two groups. CONCLUSIONS: UCP2 plays a regulatory role in the activation of p38 MAPK and NF-κB and the expression of downstream inflammatory mediators in H9C2 cells stimulated with septic serum.

Comparison of microRNA expression in hippocampus and the marginal division (MrD) of the neostriatum in rats.

BACKGROUND: MicroRNAs (miRNAs), a class of highly conserved small non-coding RNA molecules, are known to play essential roles in central nervous system (CNS) by causing post-transcriptional gene silencing. There is much evidence that miRNAs have specific temporal and spatial expression patterns in the mammal brain, but little is known about the role of the region specificity for the gene regulatory networks of the brain. This study represents the first attempt to perform a profiling analysis of the differential expression of miRNAs between hippocampus and the Marginal division (MrD) of the neostriatum in the rat brain. RESULTS: Microarray was used to detect the expression of 357 miRNAs in hippocampus and the MrD from three rats. A short-list of the most dysregulated 30 miRNAs per rat was generated for data analysis, and the miRNAs that were represented in two or three short-lists were then further analyzed. Quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was employed to validate the aberrantly expressed miRNAs obtained from the miRNA microarray analysis. A family of 11 miRNAs demonstrated differential expression between the MrD and hippocampus in more than one rat. Amongst these, miR-383 was differentially expressed in all three rats and up-regulated to the largest degree in rat one, and the ten other miRNAs, let-7d*, miR-181b, miR-187, miR-195, miR-214, miR-382, miR-411, miR-466b, miR-592 and miR-1224 were differentially expressed in at least two rats. Of these ten, besides miR-382 and miR-411 which were up-regulated in one rat and down-regulated in another, the other eight miRNAs retained a uniform direction of regulation (up-regulation or down-regulation) between different specimens. When further examined by RT-PCR, the aberrantly expressed miRNAs, except miR-383 and let-7d*, demonstrated differential expression that significantly correlated with the microarray findings. CONCLUSION: This study reported that the miRNA expression patterns in MrD was distinct from that of Hip, suggesting the role of miRNAs in the learning and memory function of the MrD probably different from hippocampus.

Bovine pulmonary surfactant alleviates inflammation and epithelial cell apoptosis in the early phase of lipopolysaccharide-induced acute lung injury in rats.

We investigate the impact of bovine pulmonary surfactant (PS) on LPS-induced ALI in vitro and in vivo to improve recognition and prevent mortality in sepsis-induced ALI. Primary alveolar type II (AT2) cells were treated with LPS alone or in combination with PS. Cell morphology observation, CCK-8 proliferation assay, flow cytometry apoptosis assay, and ELISA for inflammatory cytokine levels were performed at different time points after treatment. An LPS-induced ALI rat model was established and treated with vehicle or PS. Lung wet/dry weight ratio, histopathological changes, lung function parameters, and serum inflammatory cytokine levels were examined 6 h after PS treatment. Survival analysis by Kaplan-Meier method. RNA sequencing was conducted to identify LPS-induced differentially expressed genes in rat lungs. Proapoptotic gene expression in rat lungs was determined by Western blot. LPS significantly inhibited cell proliferation while promoting apoptosis of AT2 cells starting 2 h after treatment, accompanied by a significant increase in inflammatory cytokine production; PS reversed these effects. PS decreased the lung wet/dry ratio in septic rats, histological abnormalities, alterations in lung function parameters, and inflammatory cytokines production; while improving the overall survival of rats. LPS-induced differentially expressed genes were closely associated with apoptosis. PS attenuated LPS-induced upregulation of proapoptotic gene expression starting 2 h after treatment in AT2 cells while restoring lung ATPase activity in vivo. Bovine PS alleviates LPS-induced ALI in the early phase, possibly by suppressing inflammation and AT2 cell apoptosis, as a preemptive therapeutic agent for managing sepsis-induced ALI.

Hydrogen-rich saline regulates NLRP3 inflammasome activation in sepsis-associated encephalopathy rat model.

Sepsis-associated encephalopathy (SAE) is characterised by long-term cognitive impairment and psychiatric illness in sepsis survivors, associated with increased morbidity and mortality. There is a lack of effective therapeutics for SAE. Molecular hydrogen (H2) plays multiple roles in septic diseases by regulating neuroinflammation, reducing oxidative stress parameters, regulating signalling pathways, improving mitochondrial dysfunction, and regulating astrocyte and microglia activation. Here we report the protective effect of hydrogen-rich saline in the juvenile SAE rat model and its possible underlying mechanisms. Rats were injected intraperitoneally with lipopolysaccharide at a dose of 5 mg/kg to induce sepsis; Hydrogen-rich saline (HRS) was administered 1 h after LPS induction at a dose of 5 ml/kg and nigericin at 1 mg/kg 1 h before LPS injection. H&E staining for neuronal damage, TUNEL assay for detection of apoptotic cells, immunofluorescence, ELISA protocol for inflammatory cytokines and 8-OHdG determination and western blot analysis to determine the effect of HRS in LPS-induced septic rats. Rats treated with HRS showed decreased TNF-α and IL-1ß expression levels. HRS treatment enhanced the activities of antioxidant enzymes (SOD, CAT and GPX) and decreased MDA and MPO activities. The number of MMP-9 and NLRP3 positive immunoreactivity cells decreased in the HRS-treated group. Subsequently, GFAP, IBA-1 and CD86 immunoreactivity were reduced, and CD206 increased after HRS treatment. 8-OHdG expression was decreased in the HRS-treated rats. Western blot analysis showed decreased NLRP3, ASC, caspase-1, MMP-2/9, TLR4 and Bax protein levels after HRS treatment, while Bcl-2 expression increased after HRS treatment. These data demonstrated that HRS attenuated neuroinflammation, NLRP3 inflammasome activation, neuronal injury, and mitochondrial damage via NLRP3/Caspase-1/TLR4 signalling in the juvenile rat model, making it a potential therapeutic agent in the treatment of paediatric SAE.

Erratum - Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) ameliorates sepsis-associated acute kidney injury by maintaining mitochondrial homeostasis and improving the mitochondrial function.

This corrects the article published in European Journal of Histochemistry 2022;66:3412. doi: 10.4081/ejh.2022.3412.

Lipopolysaccharide-induced endotoxaemia during adolescence promotes stress vulnerability in adult mice via deregulation of nuclear factor erythroid 2-related factor 2 in the medial prefrontal cortex.

RATIONALE: Sepsis is a severe inflammatory response to infection that leads to long-lasting cognitive impairment and depression after resolution. The lipopolysaccharide (LPS)-induced endotoxaemia model is a well-established model of gram-negative bacterial infection and recapitulates the clinical characteristics of sepsis. However, whether LPS-induced endotoxaemia during adolescence can modulate depressive and anxiety-like behaviours in adulthood remains unclear. OBJECTIVES: To determine whether LPS-induced endotoxaemia in adolescence can modulate the stress vulnerability to depressive and anxiety-like behaviours in adulthood and explore the underlying molecular mechanisms. METHODS: Quantitative real-time PCR was used to measure inflammatory cytokine expression in the brain. A stress vulnerability model was established by exposure to subthreshold social defeat stress (SSDS), and depressive- and anxiety-like behaviours were evaluated by the social interaction test (SIT), sucrose preference test (SPT), tail suspension test (TST), force swimming test (FST), elevated plus-maze (EPM) test, and open field test (OFT). Western blotting was used to measure Nrf2 and BDNF expression levels in the brain. RESULTS: Our results showed that inflammation occurred in the brain 24 h after the induction of LPS-induced endotoxaemia at P21 but resolved in adulthood. Furthermore, LPS-induced endotoxaemia during adolescence promoted the inflammatory response and the stress vulnerability after SSDS during adulthood. Notably, the expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and BDNF in the mPFC were decreased after SSDS exposure in mice treated with LPS during adolescence. Activation of the Nrf2-BDNF signalling pathway by sulforaphane (SFN), an Nrf2 activator, ameliorated the effect of LPS-induced endotoxaemia during adolescence on stress vulnerability after SSDS during adulthood. CONCLUSIONS: Our study identified adolescence as a critical period during which LPS-induced endotoxaemia can promote stress vulnerability during adulthood and showed that this effect is mediated by impairment of Nrf2-BDNF signalling in the mPFC.

Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) ameliorates sepsis-associated acute kidney injury by maintaining mitochondrial homeostasis and improving the mitochondrial function.

Mitochondrial dysfunction has a role in sepsis-associated acute kidney injury (S-AKI), so the restoration of normal mitochondrial homeostasis may be an effective treatment strategy. Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) is a main regulator of cell-redox homeostasis, and recent studies reported that NRF2 activation helped to preserve mitochondrial morphology and function under conditions of stress. However, the role of NRF2 in the process of S-AKI is still not well understood. The present study investigated whether NRF2 regulates mitochondrial homeostasis and influences mitochondrial function in S-AKI. We demonstrated activation of NRF2 in an in vitro model: lipopolysaccharide (LPS) challenge of ductal epithelial cells of rat renal tubules (NRK-52e cells), and an in vivo model: cecal ligation and puncture (CLP) of rats. Over-expression of NRF2 attenuated oxidative stress, apoptosis, and the inflammatory response; enhanced mitophagy and mitochondrial biogenesis; and mitigated mitochondrial damage in the in vitro model. In vivo experiments showed that rats treated with an NRF2 agonist had higher adenosine triphosphate (ATP) levels, lower blood urea nitrogen and creatinine levels, fewer renal histopathological changes, and higher expression of mitophagy-related proteins [PTEN-induced putative kinase 1 (PINK1), parkin RBR E3 ubiquitin protein ligase (PRKN), microtubule-associated protein 1 light chain 3 II (LC3 II)] and mitochondrial biogenesis-related proteins [peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1α) and mitochondrial transcription factor A (TFAM)]. Electron microscopy of kidney tissues showed that mitochondrial damage was alleviated by treatment with an NRF2 agonist, and the opposite response occurred upon treatment with an NRF2 antagonist. Overall, our findings suggest that mitochondria have an important role in the pathogenesis of S-AKI, and that NRF2 activation restored mitochondrial homeostasis and function in the presence of this disease. This mitochondrial pathway has the potential to be a novel therapeutic target for the treatment of S-AKI.

Genetic Polymorphisms of LPCAT1, CHPT1 and PCYT1B and Risk of Neonatal Respiratory Distress Syndrome among a Chinese Han Population.

BACKGROUND: The study of genetic polymorphisms of surfactant-lipids related genes can help to understand individual variability in the susceptibility to development of pulmonary pathologies. The purpose of this study was to evaluate the association of polymorphisms of surfactant-lipids related genes (LPCAT1, CHPT1 and PCYT1B) with the risk/severity of respiratory distress syndrome (RDS) in preterm neonates among the Chinese Han population in Southern China. METHODS: Four hundred and forty-six preterm neonates were enrolled in a case-control study. Six polymorphisms of 3 genes were analyzed by PCR amplification of genomic DNA and genotyping was performed using an improved multiplex ligation detection reaction (iMLDR) technique based on LDR. RESULTS: The GG genotype and G allele of LPCAT1-rs9728 were found less frequently in the RDS group than in the controls (11.5% vs. 22.0% and 38.3% vs. 48.2%, respectively) (p < 0.05). CONCLUSION: This report is the first study to evaluate a direct genetic association between polymorphisms of LPCAT1 and RDS development in Chinese Han preterm infants. Our study raises the possibility that a genetic variation of LPCAT1 could be implicated in the pathophysiology of RDS in preterm neonates. GG genotype and G allele of rs9728 are protective factors for the development of RDS in preterm infants.

UCP2 ameliorates mitochondrial dysfunction, inflammation, and oxidative stress in lipopolysaccharide-induced acute kidney injury.

OBJECTIVE: UCP2 is involved in the maintenance of mitochondrial function, immune response and regulation of oxidative stress under physiological or pathological conditions. The aim of this study was to investigate the effects of UCP2 on mitochondrial dysfunction, inflammation, and oxidative stress in septic acute kidney injury (AKI). METHODS: We established LPS-induced AKI model in mice and HK-2 cells. In vivo, the UCP2 inhibitor genipin was used to downregulate UCP2 in mouse kidneys. In vitro, UCP2 overexpression or knockdown was achieved by LV5-UCP2 or si-UCP2 transfection, respectively, to characterize the mechanisms of UCP2 in septic AKI. Indicators of renal injury, cell apoptosis, inflammation, oxidative stress, and mitochondrial dysfunction were assessed. RESULTS: Compared to the control group, LPS treatment increased UCP2 expression in vitro and in vivo. In vitro, UCP2 overexpression protected HK-2 cells from LPS-induced injury by suppression of apoptosis, inflammation, oxidative stress, MMP loss and ROS production, increase of ATP production and mtDNA content, and amelioration of damage to the mitochondrial ultrastructure. Additionally, inhibition of UCP2 expression by si-UCP2 resulted in decreased HK-2 cell resistance to LPS toxicity, as shown by increased apoptosis, inflammation, mitochondrial dysfunction and oxidative stress. In vivo, UCP2 downregulation aggravated the LPS-induced renal injury, inflammation, macrophages infiltration, mitochondrial dysfunction, and oxidative stress. CONCLUSION: UCP2 may protect LPS-induced AKI by ameliorating mitochondrial dysfunction, anti-inflammation, and antioxidative activities, ultimately inhibiting tubule epithelial cell apoptosis, and that increasing the UCP2 content in mitochondria constitutes a new therapeutic approach for septic AKI.