Documenting the immune response in patients with COVID-19-induced acute respiratory distress syndrome.

Introduction: Coronavirus disease 2019 (COVID-19) can lead to acute respiratory distress syndrome (ARDS) and life-threatening multi-organ failure with increased levels of inflammatory mediators and viral load; however, little is known about its pathophysiology. Methods: To better understand the cellular status of COVID-19-induced ARDS, we performed single-cell RNA sequencing on peripheral blood samples from patients with COVID-19-induced ARDS. Single-cell RNA sequencing combined with bioinformatics analysis was used to study dynamic changes in cell composition and transcriptional profiles. Results: The single-cell RNA sequencing data revealed significant phenotypic differences between patients with COVID-19-induced ARDS and controls, mainly in monocytes, and CD8+ T and B cells. B-cell and monocyte abundances were significant in COVID-19-induced ARDS patients compared to controls, while CD8+ T cells were depleted. These data suggest that there is an imbalance between lymphocytes and monocytes in the blood of COVID-19-induced ARDS patients. In addition, cytokine interactions between T cells, monocytes and B cells are enhanced as evidenced by the intercellular communication analysis. In particular, T cell subsets target receptors on other cells via CCL5 and may play an important role in patients with COVID-19-induced ARDS. Conclusion: Our analysis suggested that a dysregulated adaptive immune response exists in patients with COVID-19-induced ARDS. Overall, we provided a cellular picture of the peripheral immune response in patients with COVID-19-induced ARDS.

[Effect of extracellular signal-regulated kinase inhibitor on calpain in rat cerebral cortex after cardiopulmonary resuscitation].

OBJECTIVE: To explore the effect of extracellular signal-regulated kinase (ERK) inhibitor PD98059 on calpain-related proteins in the brain, and to understand the pathophysiological changes of calpain in cerebral ischemia/reperfusion injury (CIRI). METHODS: Forty-two rats were divided into sham operation (Sham) group (n = 6), model group (n = 12), dimethyl sulfoxide (DMSO) control group (n = 12), and PD98059 group (n = 12) by random number table. The rat model of CIRI induced by cardiac arrest-cardiopulmonary resuscitation (CA-CPR) was reproduced by transesophageal electrical stimulation to induce ventricular fibrillation. In the Sham group, only the basic operations such as anesthesia, tracheal intubation, and arteriovenous catheterization were performed without CA-CPR. The rats in the DMSO control group and PD98059 group were injected with DMSO or PD98059 0.30 mg/kg via femoral vein, respectively, 30 minutes after the restoration of spontaneous circulation (ROSC), and rats in the Sham group and model group were given the same amount of normal saline. The duration of CPR, 24-hour survival rate and neurological deficit score (NDS) after ROSC were recorded. Hematoxylin-eosin (HE) staining and Nissl staining were used to observe the pathological changes of the cerebral cortex. The expressions of phosphorylated ERK (p-ERK), ERK, calpastatin, calpain-1, and calpain-2 were detected by Western blotting. The co-expression of p-ERK and calpain-2 was detected by double immunofluorescence. RESULTS: There were no significant differences in the duration of CPR and 24-hour survival rate among all groups. In the model group, the nuclei of the cerebral cortex were obviously deformed and pyknotic, cells vacuoles and tissues were arranged disorderly, Nissl corpuscles were significantly reduced, NDS scores were also significantly reduced, level of ERK phosphorylation was increased, and calpain-2 protein was significantly up-regulated compared with the Sham group. There was no significant difference in the above parameters between the DMSO control group and the model group. After intervention with PD98059, the pathological injury of brain tissue was significantly improved, Nissl corpuscles were significantly increased, the NDS score was significantly higher than that in the model group [75.0 (72.0, 78.0) vs. 70.0 (65.0, 72.0), P < 0.05], the level of ERK phosphorylation and calpain-2 protein expression were significantly lower than those in the model group [p-ERK (p-ERK/ERK): 0.65±0.12 vs. 0.92±0.05, calpain-2 protein (calpain-2/GAPDH): 0.73±0.10 vs. 1.07±0.14, both P < 0.05], while there was no significant difference in the expressions of calpastatin and calpain-1 in the cerebral cortex among all the groups. Double immunofluorescence staining showed that p-ERK and calpain-2 were co-expressed in cytosol and nucleus, and the co-expression rate of p-ERK and calpain-2 in the model group was significantly higher than that in the Sham group [(38.6±4.3)% vs. (9.2±3.5)%, P < 0.05], while it was significantly lowered in the PD98059 group compared with the model group [(18.2±7.0)% vs. (38.6±4.3)%, P < 0.05]. CONCLUSIONS: ERK together with calpain-2 participated in CIRI induced by CA-CPR. PD98059 inhibited the expression of calpain-2 and ERK phosphorylation. Therefore, ERK/calpain-2 may be a novel therapeutic target for CIRI.

M2c Macrophages Protect Mice from Adriamycin-Induced Nephropathy by Upregulating CD62L in Tregs.

Regulatory T cells (Tregs) and M2c macrophages have been shown to exert potentially synergistic therapeutic effects in animals with adriamycin-induced nephropathy (AN), a model chronic proteinuric renal disease. M2c macrophages may protect against renal injury by promoting an increase in the number of Tregs in the renal draining lymph nodes of AN mice, but how they do so is unclear. In this study, we used an AN mouse model to analyze how M2c macrophages induce the migration of Tregs. Using flow cytometry, we found that M2c macrophages promoted the migration of Tregs from the peripheral blood to the spleen, thymus, kidney, and renal draining lymph nodes. At the same time, M2c macrophages significantly upregulated chemokine receptors and adhesion molecule in Tregs, including CCR4, CCR5, CCR7, CXCR5, and CD62L. Treating AN mice with monoclonal anti-CD62L antibody inhibited the migration of M2c macrophages and Tregs to the spleen, thymus, kidney, and renal draining lymph nodes. Taken together, our results suggest that M2c macrophages upregulate CD62L in Tregs and thereby promote their migration to inflammatory sites, where they exert renoprotective effects. These insights may aid the development of treatments against chronic kidney disease.

Survival outcomes and adverse events in patients with chronic kidney disease after coronary artery bypass grafting and percutaneous coronary intervention: a meta-analysis of propensity score-matching studies.

BACKGROUND: The present meta-analysis of propensity score-matching studies aimed to compare the long-term survival outcomes and adverse events associated with coronary artery bypass grafting (CABG) and percutaneous coronary intervention (PCI) in patients with chronic kidney disease (CKD). METHODS: Electronic databases were searched for studies comparing CABG and PCI in patients with CKD. The search period extended to 13 February 2021. The primary outcome was all-cause mortality, and the secondary endpoints included myocardial infarction, revascularization, and stroke. Odds ratios (ORs) and hazard ratios (HRs) with 95% confidence intervals (CIs) were used to express the pooled effect. Study quality was assessed using the Newcastle-Ottawa scale. The analyses were performed using RevMan 5.3. RESULTS: Thirteen studies involving 18,005 patients were included in the meta-analysis. Long-term mortality risk was significantly lower in the CABG group than in the PCI group (HR: 0.76, 95% CI: 0.70-0.83, p < .001), and similar results were observed in the subgroup analysis of patients undergoing dialysis and for different estimated glomerular filtration rate ranges. The incidence rates of myocardial infarction (OR: 0.25, 95% CI: 0.12-0.54, p < .001) and revascularization (OR: 0.17, 95% CI: 0.08-0.35, p < .001) were lower in the CABG group than in the PCI group, although there were no significant differences in the incidence of stroke between the two groups (OR: 1.24; 95% CI: 0.89-1.73, p > .05). Subgroup analysis among patients on dialysis yielded similar results. CONCLUSIONS: Our propensity score matching analysis revealed that, based on long-term follow-up outcomes, CABG remains superior to PCI in patients with CKD.

Inhibition of extracellular signal-regulated kinase/calpain-2 pathway reduces neuroinflammation and necroptosis after cerebral ischemia-reperfusion injury in a rat model of cardiac arrest.

BACKGROUND: Cerebral ischemia-reperfusion injury (CIRI) is the leading cause of poor neurological prognosis after cardiopulmonary resuscitation (CPR). We previously reported that the extracellular signal-regulated kinase (ERK) activation mediates CIRI. Here, we explored the potential ERK/calpain-2 pathway role in CIRI using a rat model of cardiac arrest (CA). METHODS: Adult male Sprague-Dawley rats suffered from CA/CPR-induced CIRI, received saline, DMSO, PD98059 (ERK1/2 inhibitor, 0.3 mg/kg), or MDL28170 (calpain inhibitor, 3.0 mg/kg) after spontaneous circulation recovery. The survival rate and the neurological deficit score (NDS) were utilized to assess the brain function. Hematoxylin stain, Nissl staining, and transmission electron microscopy were used to evaluate the neuron injury. The expression levels of p-ERK, ERK, calpain-2, neuroinflammation-related markers (GFAP, Iba1, IL-1ß, TNF-α), and necroptosis proteins (TNFR1, RIPK1, RIPK3, p-MLKL, and MLKL) in the brain tissues were determined by western blotting and immunohistochemistry. Fluorescent multiplex immunohistochemistry was used to analyze the p-ERK, calpain-2, and RIPK3 co-expression in neurons, and RIPK3 expression levels in microglia or astrocytes. RESULTS: At 24 h after CA/CPR, the rats in the saline-treated and DMSO groups presented with injury tissue morphology, low NDS, ERK/calpain-2 pathway activation, and inflammatory cytokine and necroptosis protein over-expression in the brain tissue. After PD98059 and MDL28170 treatment, the brain function was improved, while inflammatory response and necroptosis were suppressed by ERK/calpain-2 pathway inhibition. CONCLUSION: Inflammation activation and necroptosis involved in CA/CPR-induced CIRI were regulated by the ERK/calpain-2 signaling pathway. Inhibition of that pathway can reduce neuroinflammation and necroptosis after CIRI in the CA model rats.

Cerebral protection of epigallocatechin gallate (EGCG) via preservation of mitochondrial function and ERK inhibition in a rat resuscitation model.

BACKGROUND: Various and opposite roles of epigallocatechin gallate (EGCG) have been reported in different studies. We aimed to investigate how EGCG affects the cerebral injury in a cardiac arrest/cardiopulmonary resuscitation (CA/CPR) model of rat. METHODS: The rats which were subjected to CA/CPR randomly received low dose of EGCG (3 mg/kg, Low-EGCG group, n=16), high dose of EGCG (9 mg/kg, High-EGCG group, n=16) and equal volume of 0.9% saline solution (NS group, n=16) at the first minute after return of spontaneous circulation (ROSC). The rats underwent anesthesia and intubation were defined as Sham group (n=16). Twenty-four hours after ROSC, neural defect score (NDS), ROS fluorescence intensity, degree of mitochondrial permeability transition pore (mPTP) opening, ATP contents and mitochondrial ATP synthase expression were evaluated in the four groups. The expression of extracellular signal-regulated kinase (ERK) activity and cleaved-caspase 3 were also detected by Western blot. RESULTS: CA/CPR induced severe ischemia-reperfusion injury (IRI), resulted in mitochondrial dysfunction and upregulated phosphorylation of ERK. EGCG dose-dependently alleviated the IRI after CA/CPR, inhibited ERK activity and restored mitochondrial function and, as indicated by improved NDS, reduced ROS level, decreased mPTP opening, elevated ATP content, increased ATPase expression and downregulated cleaved-caspase 3 level. CONCLUSION: EGCG alleviated global cerebral IRI by restoring mitochondrial dysfunction and ERK modulation in a rat CA/CPR model, which might make it a potential candidate agent against IRI after CA/CPR in the future. Further study is needed to determine whether higher dosage of EGCG might aggravate cerebral IRI post-CA/CPR.

High-potassium preconditioning enhances tolerance to focal cerebral ischemia-reperfusion injury through anti-apoptotic effects in male rats.

Imbalances between cellular K+ efflux and influx are considered to be involved in cerebral ischemia-reperfusion (I/R) injury. High-potassium pretreatment alleviates this injury, but the underlying molecular mechanism is unclear. In this study, we sought to investigate whether high-potassium preconditioning enhances cerebral tolerance to I/R injury through an anti-apoptotic mechanism. Adult male Sprague-Dawley rats were randomly divided into four groups (n = 40/group): a sham-operated group, normal saline group (3.2 ml/kg saline, intravenous (IV)), and low-dose and high-dose potassium chloride (KCl) groups (40 and 80 mg/kg KCl solution, IV, respectively). Subsequently, the rats underwent 90 min of middle cerebral artery occlusion (MCAO) followed by 24 hr of reperfusion (MCAO/R). Neurological deficit scores, 2,3,5-triphenyltetrazolium chloride (TTC) staining, hematoxylin and eosin staining, and TUNEL assay were used to assess neural injury. The expression of apoptotic proteins, brain potassium levels, mitochondrial function and oxidative stress were detected to explore the potential mechanism. After 24 hr of reperfusion, in both KCl treatment groups, neurological deficits and the cerebral infarct volume were reduced, and the apoptosis index of neurons was decreased. Furthermore, high-potassium preconditioning increased brain K+ , adenosine triphosphate (ATP), cytochrome c oxidase (COX) levels, reduced malondialdehyde level, improved Na+ /K+ -ATPase, succinic dehydrogenase and superoxide dismutase activities, upregulated anti-apoptotic protein expression, and downregulated pro-apoptotic protein expression. This study suggests that high-potassium preconditioning enhanced cerebral tolerance to I/R injury in a rat MCAO/R model. The protective mechanism may involve apoptosis inhibition via preservation of intracellular K+ and improvement of mitochondrial function.

The incidence of acute kidney injury following cardiac arrest and cardiopulmonary resuscitation in a rat model.

OBJECTIVE: In the current study, we investigated the incidence of acute kidney injury (AKI) induced by cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) and whether such an AKI can recover spontaneously in rats. METHODS: We used transesophageal alternating current stimulation to establish 7 min of CA rat model followed by conventional CPR. The experimental rats were randomly divided into three groups (n = 20 per group) according to the different time points after restoration spontaneous circulation (ROSC): the ROSC 24 h, ROSC 48 h, and ROSC 72 h group. The diagnosis of rat AKI refers to the 2012 KDIGO adult AKI diagnostic criteria. The severity of AKI quantified by the serum creatinine (SCR), blood urea nitrogen (BUN) levels and histological features of renal tissue. RESULTS: The incidence rates of AKI in ROSC 24 h, ROSC 48 h, and ROSC 72 h group were 65%, 45%, and 42.9%. Moreover, the values of SCR and BUN were highest at ROSC 24 h, and then gradually decreased with the time of ROSC. The histological changes of the renal tissues such as glomerular collapse, renal tubular cell swelling, and inflammatory cell infiltration had also observed. CONCLUSION: The incidence of AKI in rats was high after suffering from CA and CPR, but renal function improved with the prolongation of ROSC time, indicating the ability of the kidney to self-repair.

Elevated Serum Potassium Concentration Alleviates Cerebral Ischemia-Reperfusion Injury via Mitochondrial Preservation.

BACKGROUND/AIMS: The anti-apoptotic effect of an increase in the extracellular concentration of potassium ([K+]) has been confirmed in vitro. However, it is not yet known whether elevated serum [K+] exerts a cerebroprotective effect in vivo. In this study, we aimed to explore the effect of elevated serum [K+] in a rat model of middle cerebral artery occlusion and reperfusion (MCAO/R). METHODS: Rats subjected to 90-min MCAO received 2.5% KCL, 1.25% KCL, or a normal saline solution at a dose of 3.2 mL/kg at the onset of reperfusion. Rats that were subjected to vascular exposure and ligation without MCAO were defined as the Sham group. Serum [K+] was determined using a blood gas analyzer at 1 min after medicine administration. At 24 h post-reperfusion, rat brains were harvested and processed for 2% 2,3,5-triphenyltetrazolium chloride staining, terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate-biotin nick end labeling staining, detection of caspase-3 and cleaved-caspase-3 by western blotting, detection of reactive oxygen species (ROS) by dihydroethidium staining, and observation of mitochondrial structure by electron microscopy. In addition, malondialdehyde (MDA), adenosine triphosphate (ATP), total superoxide dismutase (T-SOD), cytochrome C oxidase (COX) activity, and mitochondrial permeability transition pore (MPTP) opening were measured using detection kits. RESULTS: The results showed that elevated serum [K+] decreased cerebral injury and apoptosis, reduced ROS and MDA levels and MPTP opening, increased ATP levels and cytochrome C oxidase activity, and improved mitochondrial ultrastructural changes, although there was no significant difference in T-SOD activity. CONCLUSION: These findings suggested that elevated serum [K+] could alleviate cerebral ischemia-reperfusion injury and the mechanism may be associated with the preservation of mitochondrial function.

Comparing the adverse clinical outcomes in patients with non-insulin treated type 2 diabetes mellitus and patients without type 2 diabetes mellitus following percutaneous coronary intervention: a systematic review and meta-analysis.

BACKGROUND: Several studies showed Type 2 Diabetes Mellitus (T2DM) to be associated with worse adverse clinical outcomes compared to non-T2DM (NDM) following Percutaneous Coronary Intervention (PCI). In addition, patients with insulin-treated T2DM (ITDM) showed worse clinical outcomes compared to patients with non-insulin treated T2DM (NITDM). Since NITDM and NDM have seldom been systematically analyzed, this study aimed to compare the short and long term adverse clinical outcomes observed in patients with NITDM and patients without T2DM following PCI. METHODS: Medline/PubMed, EMBASE and the Cochrane library were searched for Randomized Controlled Trials (RCTs) and observational studies comparing patients with (including ITDM and NITDM) and without T2DM following PCI. Endpoints included adverse clinical outcomes reported during a short and a long term follow up period. Odd Ratios (OR) and 95% Confidence Intervals (CI) in accordance with either a fixed or a random effects model appropriately, were calculated and the pooled analyses were performed with RevMan 5.3. RESULTS: Twelve studies consisting of a total number of 52,451 patients (14,863 NITDM and 37,588 NDM) were included. Patients with NITDM were found to have significantly higher short-term Major Adverse Cardiac Events (MACEs) and mortality with OR: 1.63, 95% CI (1.17, 2.27); P = 0.004 and OR: 1.71, 95% CI (1.40, 2.10), P < 0.00001 respectively and higher long-term MACEs and mortality with OR: 1.25, 95% CI (1.12, 1.40), P = 0.0001 and OR: 1.32, 95% CI (1.19, 1.47), P < 0.00001 respectively compared to NDM following PCI. In addition, compared to NDM, long-term Target Vessel Revascularization (TVR) and Target Lesion Revascularization (TLR) were significantly higher in the NITDM group with OR: 1.36, 95% CI (1.18, 1.56), P < 0.0001 and OR: 1.32, 95% CI (1.10, 1.59), P = 0.003 respectively. However, even if an increased long-term stent thrombosis was observed in the NITDM group with OR: 1.13; 95% CI (0.91, 1.40), P = 0.28, the result was insignificant. CONCLUSION: Short and long term MACEs and mortality were significantly higher in patients with NITDM compared to patients without diabetes following PCI. Revascularization also significantly favored patients without T2DM. However, stent thrombosis was not significantly different.