Identification of renal ischemia reperfusion injury-characteristic genes, pathways and immunological micro-environment features through bioinformatics approaches.

BACKGROUND: Biomarkers and pathways associated with renal ischemia reperfusion injury (IRI) had not been well unveiled. This study was intended to investigate and summarize the regulatory networks for related hub genes. Besides, the immunological micro-environment features were evaluated and the correlations between immune cells and hub genes were also explored. METHODS: GSE98622 containing mouse samples with multiple IRI stages and controls was collected from the GEO database. Differentially expressed genes (DEGs) were recognized by the R package limma, and the GO and KEGG analyses were conducted by DAVID. Gene set variation analysis (GSVA) and weighted gene coexpression network analysis (WGCNA) had been implemented to uncover changed pathways and gene modules related to IRI. Besides the known pathways such as apoptosis pathway, metabolic pathway, and cell cycle pathways, some novel pathways were also discovered to be critical in IRI. A series of novel genes associated with IRI was also dug out. An IRI mouse model was constructed to validate the results. RESULTS: The well-known IRI marker genes (Kim1 and Lcn2) and novel hub genes (Hbegf, Serpine2, Apbb1ip, Trip13, Atf3, and Ncaph) had been proved by the quantitative real-time polymerase chain reaction (qRT-PCR). Thereafter, miRNAs targeted to the dysregulated genes were predicted and the miRNA-target network was constructed. Furthermore, the immune infiltration for these samples was predicted and the results showed that macrophages infiltrated to the injured kidney to affect the tissue repair or fibrosis. Hub genes were significantly positively or negatively correlated with the macrophage abundance indicating they played a crucial role in macrophage infiltration. CONCLUSIONS: Consequently, the pathways, hub genes, miRNAs, and the immune microenvironment may explain the mechanism of IRI and might be the potential targets for IRI treatments.

Exosomes derived from bone marrow mesenchymal stem cells regulate pyroptosis via the miR-143-3p/myeloid differentiation factor 88 axis to ameliorate intestinal ischemia-reperfusion injury.

Intestinal ischemia-reperfusion (I/R) injury is a condition in which tissue injury is aggravated after ischemia due to recovery of blood supply. Bone marrow mesenchymal stem cell-derived exosome (BMSC-exo) showed a protective effect on I/R injury. This study aimed to investigate the possible mechanisms by which BMSC-exos ameliorate intestinal I/R injury. We isolated mouse BMSC-exos by super-centrifugation and found that they effectively increased cell viability in a cell model, alleviated intestinal barrier injury in a mouse model, and downregulated the expression of inflammatory cytokines and pyroptosis-related proteins, suggesting that BMSC-exos may alleviate intestinal I/R injury in vitro and in vivo by regulating pyroptosis. We identified miR-143-3p as a differentially expressed miRNA by microarray sequencing. Bioinformatic analysis predicted a binding site between miR-143-3p and myeloid differentiation factor 88 (MyD88); a dual-luciferase reporter assay confirmed that miR-143-3p could directly regulate the expression of MyD88. Our findings suggest that miR-143-3p regulates pyroptosis by regulating NOD-like receptor thermal protein domain associated protein 3 (NLRP3) through the toll-like receptor (TLR)-4/MyD88/nuclear factor kappa-B (NF-кB) pathway. This study describes a potential strategy for the treatment of intestinal I/R injury using BMSC-exos that act by regulating pyroptosis through the miR-143-3p mediated TLR4/MyD88/NF-кB pathway.

BMSC-exos ameliorate intestinal ischemia/reperfusion (I/R) injurymiR-143-3p levels were reduced in I/R injury and increased with BMSC-exo treatmentmiR-143-3p directly targeted and downregulated the expression of MyD88BMSC-exos regulate pyroptosis in intestinal I/R injury via the miR-143-3p-MyD88 axis.

miRNA155-5P participated in DDX3X targeted regulation of pyroptosis to attenuate renal ischemia/reperfusion injury.

BACKGROUND: Renal ischemia/reperfusion injury (IRI) induced pathological damage to renal microvessels and tubular epithelial cells through multiple factors. However, studies investigated whether miRNA155-5P targeted DDX3X to attenuate pyroptosis were scarce. RESULTS: The expression of pyroptosis-related proteins (caspase-1, interleukin-1ß (IL-1ß), NOD-like receptor family pyrin domain containing 3 (NLRP3), and IL-18) were up-regulated in the IRI group. Additionally, miR-155-5p was higher in the IRI group comparing with the sham group. The DDX3X was inhibited by the miR-155-5p mimic more than in the other groups. DEAD-box Helicase 3 X-Linked (DDX3X), NLRP3, caspase-1, IL-1ß, IL-18, LDH, and pyroptosis rates were higher in all H/R groups than in the control group. These indicators were higher in the miR-155-5p mimic group than in the H/R and the miR-155-5p mimic negative control (NC) group. CONCLUSIONS: Current findings suggested that miR-155-5p decreased the inflammation involved in pyroptosis by downregulating the DDX3X/NLRP3/caspase-1 pathway. METHODS: Using the models of IRI in mouse and the hypoxia-reoxygenation (H/R)-induced injury in human renal proximal tubular epithelial cells (HK-2 cells), we analyzed the changes in renal pathology and the expression of factors correlated with pyroptosis and DDX3X. Real-time reverse transcription polymerase chain reaction (RT-PCR) detected miRNAs and enzyme-linked immunosorbent assay (ELISA) was used to detect lactic dehydrogenase activity. The StarBase and luciferase assays examined the specific interplay of DDX3X and miRNA155-5P. In the IRI group, severe renal tissue damage, swelling, and inflammation were examined.

Role of microRNAs in programmed cell death in renal diseases: A review.

MicroRNAs (miRNAs) regulate gene expression involving kidney morphogenesis and cell proliferation, apoptosis, differentiation, migration, invasion, immune evasion, and extracellular matrix remodeling. Programmed cell death (PCD) is mediated and regulated by specific genes and a wealth of miRNAs, which participate in various pathological processes. Dysregulation of miRNAs can disrupt renal development and induce the onset and progression of various renal diseases. An in-depth understanding of how miRNAs regulate renal development and diseases is indispensable to comprehending how they can be used in new diagnostic and therapeutic approaches. However, the mechanisms are still insufficiently investigated. Hence, we review the current roles of miRNA-related signaling pathways and recent advances in PCD research and aim to display the potential crosstalk between miRNAs and PCD. The prospects of miRNAs as novel biomarkers and therapeutic targets are also described, which might provide some novel ideas for further studies.

Mesenchymal stem cells against intestinal ischemia-reperfusion injury: a systematic review and meta-analysis of preclinical studies.

BACKGROUND: Intestinal ischemia-reperfusion injury (IRI) causes localized and distant tissue lesions. Multiple organ failure is a common complication of severe intestinal IRI, leading to its high rates of morbidity and mortality. Thus far, this is poorly treated, and there is an urgent need for new more efficacious treatments. This study evaluated the beneficial effects of mesenchymal stem cells (MSCs) therapy on intestinal IRI using many animal experiments. METHODS: We conducted a comprehensive literature search from 4 databases: Pubmed, Embase, Cochrane library, and Web of science. Primary outcomes included the survival rate, Chiu's score, intestinal levels of IL-6, TNF-α and MDA, as well as serum levels of DAO, D-Lactate, and TNF-α. Statistical analysis was carried out using Review Manager 5.3. RESULTS: It included Eighteen eligible researches in the final analysis. We demonstrated that survival rates in animals following intestinal IRI were higher with MSCs treatment compared to vehicle treatment. Besides, MSCs treatment attenuated intestinal injury caused by IRI, characterized by lower Chiu's score (- 1.96, 95% CI - 2.72 to - 1.19, P < 0.00001), less intestinal inflammation (IL-6 (- 2.73, 95% CI - 4.19 to - 1.27, P = 0.0002), TNF-α (- 3.00, 95% CI - 4.74 to - 1.26, P = 0.0007)) and oxidative stress (MDA (- 2.18, 95% CI - 3.17 to - 1.19, P < 0.0001)), and decreased serum levels of DAO (- 1.39, 95% CI - 2.07 to - 0.72, P < 0.0001), D-Lactate (- 1.54, 95% CI - 2.18 to - 0.90, P < 0.00001) and TNF-α (- 2.42, 95% CI - 3.45 to - 1.40, P < 0.00001). The possible mechanism for MSCs to treat intestinal IRI might be through reducing inflammation, alleviating oxidative stress, as well as inhibiting the apoptosis and pyroptosis of the intestinal epithelial cells. CONCLUSIONS: Taken together, these studies revealed that MSCs as a promising new treatment for intestinal IRI, and the mechanism of which may be associated with inflammation, oxidative stress, apoptosis, and pyroptosis. However, further studies will be required to confirm these findings.

Exosomes Derived from BMSCs Ameliorate Intestinal Ischemia-Reperfusion Injury by Regulating miR-144-3p-Mediated Oxidative Stress.

BACKGROUND: Intestinal ischemia-reperfusion (I/R) injury is a critical pathophysiological process involved in many acute and critical diseases, and it may seriously threaten the lives of patients. Exosomes derived from bone marrow mesenchymal stem cells (BMSC-exos) may be an effective therapeutic approach for I/R injury. AIMS: This study aimed to investigate the role and possible mechanism of BMSC-exos in intestinal I/R injury in vivo and in vitro based on the miR-144-3p and PTEN/Akt/Nrf2 pathways. METHODS: BMSC-exos were isolated from mouse BMSCs by super centrifugation methods. The effects of BMSC-exos on I/R intestinal injury, intestinal cell apoptosis, oxidative stress and the PTEN/Akt/Nrf2 pathway were explored in vivo and in vitro. Furthermore, the relationship between miR-144-3p and PTEN was confirmed by a dual-luciferase reporter assay. The miR-144-3p mimic and inhibitor were used to further clarify the role of miR-144-3p in the mitigation of intestinal I/R by BMSC-exos. RESULTS: BMSC-exos effectively alleviated intestinal pathological injury, reduced intestinal cell apoptosis, relieved oxidative stress and regulated the PTEN/Akt/Nrf2 pathway in vivo and in vitro. In addition, miR-144-3p was significantly reduced in the oxygen and glucose deprivation/reperfusion cell model, and miR-144-3p could directly target PTEN to regulate its expression. Additional studies showed that changing the expression of miR-144-3p in BMSC-exos significantly affected the regulation of intestinal injury, intestinal cell apoptosis, oxidative stress and the PTEN/Akt/Nrf2 pathway in I/R, suggesting that miR-144-3p in BMSC-exos plays an important role in regulating the PTEN/Akt/Nrf2 during intestinal I/R. CONCLUSIONS: BMSC-exos carrying miR-144-3p alleviated intestinal I/R injury by regulating oxidative stress.

Dexmedetomidine Relieves Neuropathic Pain in Rats With Chronic Constriction Injury via the Keap1-Nrf2 Pathway.

This study aimed to determine the role of dexmedetomidine (Dex) in neuropathic pain (NP) after chronic constriction injury (CCI) in a rat model as well as its underlying mechanism. First, a CCI rat model was established. After treatment with Dex, the severity of NP was ascertained by monitoring paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) at different time points. Immunohistochemical analysis was performed to determine the levels of Keap1 and Nrf2 in the spinal cord. Furthermore, the levels of Keap1-Nrf2-HO-1 pathway molecules, apoptotic proteins, and antioxidant genes in the spinal cord or isolated primary microglia were determined using quantitative polymerase chain reaction and western blotting. The release of proinflammatory cytokines was detected via enzyme-linked immunosorbent assay. To evaluate Dex-treated CCI-induced NP via the Keap1-Nrf2-HO-1 pathway, the rats were intrathecally injected with lentivirus to upregulate or downregulate the expression of Keap1. We found that Dex inhibited pathological changes and alleviated sciatic nerve pain as well as repressed inflammation, apoptosis, and redox disorders of the spinal cord in CCI rats. Keap1 protein expression was substantially downregulated, whereas Nrf2 and HO-1 expressions were significantly upregulated in the spinal cord after Dex administration. Additionally, Keap1 overexpression counteracted Dex-mediated inhibition of NP. Keap1 overexpression led to a decrease in Nrf2 and HO-1 levels as well as PWT and PWL but led to an aggravation of inflammation and antioxidant disorders and increased apoptosis. Keap1 silencing alleviated NP in rats with CCI, as evidenced by an increase in PWT and PWL. Keap1 depletion resulted in the alleviation of inflammation and spinal cord tissue injury in CCI rats. Collectively, these findings suggest that Dex inhibits the Keap1-Nrf2-HO-1-related antioxidant response, inflammation, and apoptosis, thereby alleviating NP in CCI rats.

Efficacy and safety of ibuprofen in children with musculoskeletal injuries: A systematic review and meta-analysis of randomized controlled trials.

OBJECTIVE: To evaluate the analgesic efficacy and safety of ibuprofen in children with musculoskeletal injuries. METHODS: PubMed, EMBASE, Web of science, and the Cochrane Central register of Controlled Trials (CENTRAL) were systematically searched to identify eligible randomized controlled trials (RCTs) that compared ibuprofen with other analgesics for pain relief in children with musculoskeletal injuries. Primary outcomes included change of visual analog scale (VAS) scores from baseline to post-medication, the proportion of patients achieving adequate analgesia, and the proportion of patients requiring additional analgesia. Secondary outcome was the incidence of adverse effects. Data analysis was performed using RevMan 5.3 software. RESULTS: Five RCTs involving 1034 patients were included in this meta-analysis. Compared to the control group, change of VAS scores was greater in ibuprofen group at 60 min (standardized mean difference [SMD] = 0.28; 95% confidence intervals [CI], 0 to 0.57; P = .05), 90 min (SMD = 0.38; 95% CI, 0.17 to 0.59; P = .0005), and 120 min (SMD = 0.4; 95% CI, 0.23 to 0.57; P < .00001) after treatment. No difference was found in the change of VAS scores at 30 min (SMD = 0.07; 95% CI, -0.08 to 0.22; P = .36) after treatment. The proportion of patients who received adequate analgesia was higher in the ibuprofen group (risk ratios [RR] = 1.36; 95% CI, 1.20 to 1.56; P < .00001). The proportion of patients that required additional analgesia was lower in the ibuprofen group (RR = 0.7; 95% CI, 0.53 to 0.92; P = .01). The incidence of total adverse effects was lower in the ibuprofen group (RR = 0.59; 95% CI, 0.45 to 0.79; P = .0002). CONCLUSIONS: Ibuprofen provides a better pain relief with a lower incidence of adverse effects in children with musculoskeletal injuries as compared to other analgesics. PROSPERO REGISTRATION NUMBER: CRD42021231975.

Hippocampal astrocyte dysfunction contributes to etomidate-induced long-lasting synaptic inhibition.

Cognitive impairments following the use of general anesthetics are well documented but the underlying mechanisms are unclear. Here, long-lasting cognitive deficits were observed in aged mice following administration of etomidate at a clinically relevant concentration (20 mg/kg); these deficits were closely related to hippocampal synaptic inhibition and astrocyte dysfunction. Using microdialysis and magnetic-activated cell-sorting techniques, we found that astrocyte secretion of glutamate, d-serine, and ATP, as well as astrocyte function, were depressed in the hippocampus following treatment with etomidate. Interestingly, hippocampal astrocyte inhibition (designer receptors exclusively activated by designer drugs; DREADDs) had no effect on the initial synaptic inhibition, but reversed synaptic and cognitive depression in the long term. Furthermore, continual activation of hippocampal astrocytes following administration of a sedative dose (8 mg/kg) of etomidate induced synaptic inhibition and cognitive dysfunction. Our results indicate that general anesthetic-induced hippocampal astrocyte dysfunction plays a role in maintaining synaptic inhibition, which eventually induces long-lasting cognitive deficits.

Dexmedetomidine Relieves Acute Inflammatory Visceral Pain in Rats through the ERK Pathway, Toll-Like Receptor Signaling, and TRPV1 Channel.

Dexmedetomidine (DEX) is a highly selective α2 adrenergic receptor agonist. In this study, we aimed to characterize the antinociceptive effects of DEX in acute inflammatory visceral pain (AIVP) induced by acetic acid in rats and to evaluate whether antinociception was regulated by the extracellular signal-regulated protein kinase (ERK) pathway, Toll-like receptor (TLR) signaling, and transient receptor potential (TRP) channel. Acetic acid was administered to 30 male rats with or without DEX. Rats were divided into six groups, as follows: control, disease (received no treatment before acetic acid administration), vehicle-treated, low-dose DEX (lDEX), medium-dose DEX (mDEX), and high-dose DEX (hDEX)-treated groups. Thermal withdrawal latency (TWL), mechanical withdrawal threshold (MWT), and abdominal withdrawal reflex (AWR) were measured to assess pain. We detected electromyographic (EGM) responses in the rectus abdominis muscle and measured the average arterial blood pressure. Levels of interleukin 1 (IL-1), IL-2, and IL-6 in the serum, as well as tumor necrosis factor α (TNF-α) and prostaglandin E2 (PGE2) in the peritoneal fluid, were measured by ELISA. The expression levels of phospho(p)CREB, pERK1/2, pMEK1, and TRP cation channel subfamily V member 1 (TRPV1), as well as the activation state of TLR4, were determined in the spinal cord of rats by real-time polymerase chain reaction and western blot analysis. TWL and MWT scores were elevated (P < 0.05) in the hDEX and mDEX groups, whereas AWR scores decreased (P < 0.01), compared to those in the disease group. The medium and high doses of DEX suppressed IL-1, IL-6, TNF-α, and PGE2 release, and increased IL-2 release. In addition, protein and mRNA levels of MEK, ERK, and CREB were reduced in the mDEX and hDEX groups. Moreover, TLR4 and its downstream target, nuclear factor kappa B, along with calcitonin gene-related peptide release through the TRPV1 channel, were suppressed by mDEX and hDEX treatment. Taken together, our results suggest that DEX might exert an antinociceptive effect in AIVP in rats through the MEK/ERK pathway, TLR signaling, and TRPV1 channel, resulting in suppression of visceral hypersensitivity.

Dexmedetomidine relieves formaldehyde-induced pain in rats through both α2 adrenoceptor and imidazoline receptor.

Dexmedetomidine (DEX) is a highly selective α2 adrenergic receptor agonist. In this study, we investigated the analgesic effect and the underlying mechanisms of DEX on inflammatory visceral pain in rats. Twenty-five male Sprague Dawley (SD) rats were randomly divided into 5 groups, including control, sham, low dose DEX, medium dose DEX and high dose DEX group. Pain was induced with 10% formalin and scored every 15min till 2 h-post the induction. Hematoxylin-eosin (HE) staining was used to evaluate the toxicity of DEX on spinal cord neurons. Acetycholine (Ach) and noradrenaline (NA) levels were determined by using ELISA method. The expressions of natural nitric oxide synthase (nNOS), protein kinase γ (PKCγ) and protease-activated receptor 2 (PAR2) were determined by using western blot. DEX treatment relieved formaldehyde-induced pain in rats in a dose-dependent manner. Furthermore, DEX showed little neuro-toxicity on the spinal cord neurons, even at the highest dosage used in our study. Ach level was significantly increased in Sham group compared with control group. DEX treatment decreased NA levels and increased Ach levels in the incubation medium of spinal cord sections. Western blot analysis showed that the expression of nNOS, PKCγ and PAR2 was significantly decreased in DEX group compared with Sham group, whereas these effects of DEX on nNOS, PKCγ and PAR2 were blocked by both yohimbine and idazoxan, indicating that the analgesic effect of DEX is mediated by both α2 adrenergic receptor and imidazoline receptor. Yohimbine and idazoxan treatment significantly enhanced pain scores compared to DEX group, and which antagonizes the effects DEX. In conclusion, our study demonstrated that DEX could inhibit formaldehyde-induced pain by inhibiting nNOS, PKCγ and PAR2 expression through α2 adrenergic receptor and imidazoline receptor.

Protective effect of Shenfu injection preconditioning on lung ischemia-reperfusion injury.

Lung ischemia-reperfusion injury remains a problem in thoracic surgery, as minimal progress has been made concerning its prevention and control. In the present study, the protective effects and the underlying mechanism of Shenfu injection preconditioning on a rat lung ischemia-reperfusion model was investigated. Shenfu injection is a well-known Chinese traditional medicine, which is composed of Red Radix Ginseng and Radix Aconitum carmichaelii, with ginseng saponin and aconitum alkaloids as the active ingredients. A total of 72 specific pathogen-free, healthy male Wistar rats were randomly divided into control, model and Shenfu injection (10 ml/kg injection prior to injury) groups and were assessed at the following points: Ischemia 45 min; reperfusion 60 min; and reperfusion 120 min. Blood collected from the aorta abdominalis was cryopreserved at -70°C for the analysis of malondialdehyde (MDA) and superoxide dismutase (SOD) activity. Lung tissues were divided into three equal sections in order to assess the wet-to-dry (W/D) lung ratio, tumor necrosis factor (TNF)-α expression levels, myeloperoxidase (MPO) activity, alveolar damage, total protein and hematoxylin and eosin staining. The results demonstrated that the lung W/D weight ratio, TNF-α expression levels and SOD activity in the Shenfu group were significantly lower at 120 min reperfusion (P<0.05), as compared with the model group. MPO and MDA activity significantly decreased following reperfusion at 60 and 120 min (P<0.05), as compared with the model group. In addition, the degree of alveolar damage in the Shenfu group was significantly decreased (P<0.05), as compared with the model group. In addition, compared with the model group, the degree of alveolar damage in the Shenfu group was significantly lower (P<0.05); however, no significant changes in total protein were observed. The extent of alveolar structural damage and the proportion of interstitial neutrophils and alveolar and interstitial red blood cells were lower in the Shenfu group, as compared with the model and control groups. Therefore, the results of the present study suggested that Shenfu injection may have protective effects on lung ischemia-reperfusion injury.

The effects of neuromuscular blockade on operating conditions during general anesthesia for spinal surgery.

BACKGROUND: Muscle relaxants are prescribed routinely for patients undergoing general anesthesia, but the requirement for paralysis in spinal surgery is unclear. This study compared the operating conditions of general anesthesia with and without a muscle relaxant on spinal surgery patients. METHODS: Eighty-six adults who underwent elective spinal surgery were randomly assigned to a relaxant group (group R) or a no-relaxant group (group NR). All patients were induced with intravenous midazolam (0.05 mg/kg), fentanyl (4 µg/kg), propofol (1.0 mg/kg), and succinylcholine (2 mg/kg) and then atracurium was used in group R but not in group NR. The operating conditions, including muscle tone, body movements, airway pressure, anesthetics consumption, eye-opening time, extubation time, and the Observer's Assessment of the Alertness/Sedation (OAA/S) score 20 minutes after the extubation were compared between the 2 groups. RESULTS: The operating conditions including muscle tone scales, body movements, and airway pressure did not differ between the 2 groups. Eye-opening time (9.35±2.34 vs. 11.02±2.50 min; P=0.002) and extubation time (13.95±3.41 vs. 16.72±3.67 min; P=0.001) were shorter in group NR than in group R. The BIS score at extubation (87.2±5.0 vs. 83.3±5.7; P=0.001) and the OAA/S score 20 minutes after extubation (5 [3 to 5] vs. 4 [3 to 5]; P=0.005) were significantly higher in group NR than in group R. Propofol consumption was higher in group NR than in group R (4206.10±415.80 vs. 3900.60±365.40 µg/kg, respectively; P=0.001). CONCLUSIONS: General anesthesia without muscle relaxant provides similar working conditions to those observed with muscle relaxant, and it is associated with earlier eye opening and extubation and higher level of consciousness on emergence from spinal surgery.