Sevoflurane blocks KLF5-mediated transcriptional activation of ITGB2 to inhibit macrophage infiltration in hepatic ischemia/reperfusion injury.

BACKGROUND: Sevoflurane (Sevo) preconditioning and postconditioning play a protective role against injury induced by hepatic ischemia/reperfusion (I/R). At the same time, the involvement of macrophage infiltration in this process and the precise mechanisms are unclear. Here, we designed this research to elucidate the protective effects of Sevo against hepatic I/R injury and the molecules involved. METHODS: The alleviating effect of Sevo on the liver injury was analyzed by liver function analysis, hematoxylin and eosin staining, Masson trichrome staining, terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling, western blot analysis and an enzyme-linked immunosorbent assay. An in vitro cell model was developed using alpha mouse liver 12 (AML12) cells, and the cell model was treated with oxygen-glucose deprivation and reoxygenation and Sevo. Multiple bioinformatics databases were used to screen transcriptional regulators related to hepatic I/R injury and the targets of Krueppel-like factor 5 (KLF5). KLF5 expression was artificially upregulated alone or with integrin beta-2 (ITGB2) knockdown to substantiate their involvement in Sevo-mediated hepatoprotection. RESULTS: Sevo protected the liver against I/R injury by reducing cell apoptosis and inflammatory response. KLF5 was upregulated in liver tissues following I/R injury, whereas KLF5 overexpression aggravated macrophage infiltration and liver injury induced by I/R injury. KLF5 bound to the promoter of ITGB2 to enhance ITGB2 transcription. Knockdown of ITGB2 reversed the aggravation of injury caused by KLF5 overexpression in mice and AML12 cells. CONCLUSIONS: Sevo blocked KLF5-mediated transcriptional activation of ITGB2, thereby inhibiting macrophage infiltration in hepatic I/R injury.

FBXW7 promotes pathological cardiac hypertrophy by targeting EZH2-SIX1 signaling.

F-box and WD repeat domain-containing 7 (FBXW7) is an E3-ubiquitin ligase, which serves as one of the components of the SKP1, CUL1, and F-box protein type ubiquitin ligase (SCF) complex. Previous studies reveal that FBXW7 participates in cancer, inflammation and Parkinson's disease. FBXW7 also contributes to angiogenesis of endothelial cells. However, the function of FBXW7 in cardiac homeostasis remains to elucidate. Here we identified the critical role of FBXW7 during cardiac hypertrophy in humans and rodents. Quantitative real-time PCR (qRT-PCR) and Western blot revealed that the mRNA and protein levels of FBXW7 were upregulated significantly in hypertrophic hearts in human and mouse as well as Angiotensin II (Ang II)-induced hypertrophic neonatal rat cardiomyocytes (NRCM). Gain-of-function (adenovirus) and loss-of-function (siRNA) experiments provided evidence that FBXW7 promoted Ang II-induced cardiomyocyte hypertrophy as demonstrated by the increase in the size of cardiomyocytes and overexpression of hypertrophic fetal genes myosin heavy chain 7 (Myh7) natriuretic peptide a (Nppa), brain natriuretic peptide (Nppb). Further mechanism study revealed that FBXW7 promoted the expression of sine oculis homeobox homolog 1 (SIX1) in cardiomyocytes, which relied on regulation of the stability of the histone methyltransferase EZH2 (Enhancer of zeste homolog 2). Previous work revealed the pro-hypertrophic role of the EZH2-SIX1 axis in rodents. Indeed, our genetic and pharmacological evidence showed that the EZH2-SIX1 signaling was critically involved in FBXW7 functions in Ang II-induced cardiomyocyte hypertrophy. Therefore, we identified FBWX7 as an important regulator of cardiac hypertrophy via modulating the EZH2-SIX1 axis.

Glycine-nitronyl nitroxide conjugate protects human umbilical vein endothelial cells against hypoxia/reoxygenation injury via multiple mechanisms and ameliorates hind limb ischemia/reperfusion injury in rats.

Oxidative stress and inflammation play important roles in the pathogenesis of ischemia/reperfusion (I/R)-injury. The administration of antioxidants and anti-inflammatory agents has been applied to prevent I/R-injury for several decades. Of the numerous compounds administrated therapeutically in anti-oxidative stress, nitronyl nitroxide has gained increasing attention due to its continuous ability to scavenge active oxygen radicals. However, its effect is not ideal in clinical therapy. In previous study, we linked the anti-inflammatory amino acid glycine to nitronyl nitroxide and developed a novel glycine-nitronyl nitroxide (GNN) conjugate, which showed a synergetic protection against renal ischemia/reperfusion injury. However, the underlying mechanism remains unclear. In this study, a hypoxia/reoxygenation (H/R) injury model was established in human umbilical vein endothelial cells (HUVECs) and we found that the GNN conjugate significantly elevated the cell viability via reducing the apoptosis rate in H/R-treated HUVECs. Meanwhile, GNN conjugate attenuated H/R induced mitochondrial fragmentation, mitochondrial membrane potential reduction, Cytochrome c release and autophagy. To determine the extensive applicability of GNN conjugate in different I/R models and its effect in remote organs, an in vivo hind limb I/R model was established. As expected, GNN conjugate ameliorated damages of muscle and remote organs. These results demonstrate that GNN conjugate may be an effective agent against ischemia/reperfusion injury in clinical therapy.

Sevoflurane Preconditioning Downregulates GRIA1 Expression to Attenuate Cerebral Ischemia-Reperfusion-Induced Neuronal Injury.

Cerebral ischemia/reperfusion (I/R) injury is the main cause of death following trauma. The neuroprotective effect of sevoflurane (Sev) has been implicated in cerebral I/R injury. However, the mechanisms remain elusive. In this study, we aimed to explore its function in PC12 exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) and in rats challenged with I/R. Sev pretreatment reduced the damage of PC12 cells after OGD/R treatment. Moreover, Sev pretreatment ameliorated neurobehavioral deficits induced by I/R treatment, reduced brain infarct volume, and decreased apoptosis of neurons in hippocampal tissues. Sev pretreatment reduced the surface expression of glutamate receptor 1 (GRIA1) in neurons, while GRIA1 reduced the neuroprotective effects of Sev pretreatment in vitro and in vivo. There was no difference in the surface expression of GRIA2 in rats with I/R and PC12 cells exposed to OGD/R. The ratio of GRIA1/GRIA2 surface expression was reduced, and calcium permeable-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (CP-AMPAR) was blocked by Sev. Together, Sev might exert beneficial effects on cerebral I/R-induced neuronal injury through inhibiting the surface expression of GRIA1 and blocking CP-AMPAR.

Surgical timing of endoluminal repair of Stanford type B aortic coarctation and relationship to prognosis: a single-center retrospective cohort study.

Background: Stanford type B aortic dissection (TBAD) is a rare cardiovascular emergency with rapid onset and great harm. Currently, no relevant studies have analyzed the difference in clinical benefits of endovascular repair in patients with TBAD in acute and non-acute stages. To investigate the clinical characteristics and prognosis of endovascular repair in patients with TBAD at different surgical timing. Methods: The medical records of 110 patients with TBAD from June 2014 to June 2022 were retrospectively selected as the study subjects. The patients were divided into an acute group (onset time ≤14 days) and a non-acute group (onset time >14 days) according to the time to surgery, and the two groups were compared in terms of surgery and hospitalization, aortic remodeling, and follow-up results. Univariate and multivariate logistic regression were used to analyze the factors affecting the prognosis of TBAD treated with endoluminal repair. Results: The proportion of pleural effusion, heart rate, the rate of complete thrombosis of the false lumen and the difference in the maximum diameter of the false lumen in the acute group were higher than those in the non-acute group (P=0.015, <0.001, 0.029, <0.001). The length of hospital stay and the maximum postoperative diameter of the false lumen was lower than in the non-acute group (P=0.001, 0.004). There was no statistically significant difference between the two groups in the technical success rate, overlapping stent length, overlapping stent diameter, immediate postoperative contrast type I endoleak, incidence of renal failure, ischemic disease, endoleaks, aortic dilatation, retrograde type A aortic coarctation, and death (P=0.386, 0.551, 0.093, 0.176, 0.223, 0.739, 0.085, 0.098, 0.395, 0.386); coronary artery disease [odds ratio (OR) =6.630, P=0.012], pleural effusion (OR =5.026, P=0.009), non-acute surgery (OR =2.899, P=0.037), and involvement of the abdominal aorta (OR =11.362, P=0.001) were all independent risk factors affecting the prognosis of TBAD treated with endoluminal repair. Conclusions: Acute phase endoluminal repair of TBAD may contribute to aortic remodeling, and the prognosis of TBAD patients can be assessed clinically in combination with coronary artery disease, pleural effusion, and involvement of the abdominal aorta for early intervention to reduce the associated mortality.

[Network visual data mining and analysis of literature on electroacupuncture treatment of post-stroke sequelae].

OBJECTIVE: To analyze the optimum stimulating parameters and acupoint combination law of electroacupuncture (EA) treatment of post-stroke sequelae (PSS) through the network visual data mining and analysis of the literature，so as to provide reference for clinical application. METHODS: The related articles of acupuncture treatment of PSS patients published from January 1, 2005 to December 31, 2021 were collected from databases of CNKI, Wanfang, Weipu（VIP), SinoMed, CHAOXING, DUXIU, PubMed, Ovid, Embase and Cochrane Library. After the articles were screened according to our formulated inclusion and exclusion criteria, a "Clinical Literature Database of EA Treatment of PSS" was established. The SPSS Modeler14.1 software was used for modeling (by using the parameters of EA, types of apoplexy sequelae, names of acupoints, methods of acupoint matching, acupoint-attributed meridians, etc.) and association rule analysis, and the Cytoscape 3.6.0 software was used for data network visualization analysis. RESULTS: A total of 354 articles were collected, including 13 types of PSS, among which the hemi-plegia accounts for the highest proportion (51.13%), followed by depression (17.51%) and dysphagia (14.97%). A total of 228 acupoints are used, with a total frequency-time of 1 690. Among the 8 PSS (hemiplegia, depression, dysphagia, cognitive impairment, urinary incontinence, urinary retention, speech disorders, constipation) with the frequency of application of acupoints greater than or equal to 3, 275 articles are involved, with the frequency of dense wave being the highest (123 times), followed by continuous waves (95 times). The stimulation frequency of EA appears 275 times, with the low frequency being the highest (188 times). For hemiplegia type of PSS, low frequency and continuous waves or dense waves are mostly used, and the high correlation acupoint groups are Zusanli(ST36)/Hegu(LI4), LI4/Quchi(LI11) and ST36/LI11. For depression, low frequency and dense waves are mostly used, and the high correlation acupoint groups are Baihui(GV20)/Yintang(EX-HN3), GV20/Shenting(GV24) and GV20/Neiguan(PC6). For swallowing disorder, the mostly used stimulation parameter is low frequency, and the high correlation acupoint groups are Lianquan(CV23)/ST36/Fenglong(ST40), Tongli(HT5)/Yifeng(TE17)/PC6/ST40, and TE17/HT5/PC6/LI4. CONCLUSION: EA therapy is frequently used in the treatment of PSS, for which ST36, LI4 and LI11 are most frequently used in the treatment of hemiplegia, with the stimulation parameters being low-frequency and continuous waves or dense waves. For depression, GV20, EX-HN3, GV24 and PC6 are used, with the stimulation parameters being low frequency and dense waves. For dysphagia, CV23, HT5, TE17, PC6, ST36, ST40 and LI4 are employed, with low frequency electrical stimulation.

Sulforaphane Ameliorates Limb Ischemia/Reperfusion-Induced Muscular Injury in Mice by Inhibiting Pyroptosis and Autophagy via the Nrf2-ARE Pathway.

Background: Limb ischemia/reperfusion (I/R) injury, as a life-threatening syndrome, is commonly caused by skeletal muscle damage resulting from oxidative stress. Additionally, inflammation-induced pyroptosis and dysregulated autophagy are vital factors contributing to the aggravation of I/R injury. Of note, sulforaphane (SFN) is a natural antioxidant, but whether it worked in limb I/R injury and the possible mechanism behind its protection for skeletal muscle has not been clearly established. Methods: Effects of SFN on limb I/R-injured skeletal muscle were assessed by HE staining, followed by assessment of wet weight/dry weight (W/D) ratio of muscle tissues. Next, ELISA and biochemical tests were used to measure the inflammatory cytokine production and oxidative stress. Immunofluorescent analysis and Western blot were adopted to examine the level of pyroptosis- and autophagy-related proteins in vivo. Moreover, protein levels of Nrf2-ARE pathway-related factors were also examined using Western blot. Results: SFN treatment could protect skeletal muscle against limb I/R injury, as evidenced by diminished inflammation, pyroptosis, autophagy, and oxidative stress in skeletal muscles of mice. Further mechanistic exploration confirmed that antioxidative protection of SFN was associated with the Nrf2-ARE pathway activation. Conclusions: SFN activates the Nrf2-ARE pathway, and thereby inhibits pyroptosis and autophagy and provides a novel therapeutic strategy for the limb I/R-induced muscle tissue damage.

Dl-3-n-Butylphthalide (NBP) Mitigates Muscular Injury Induced by Limb Ischemia/Reperfusion in Mice through the HMGB1/TLR4/NF-κB Pathway.

Objective: Limb ischemia/reperfusion (I/R) injury is a clinical syndrome associated with severe damages to skeletal muscles and other fatal outcomes. Oxidative stress and inflammatory response play vital roles in the development of limb I/R injury. Existing evidence further indicates that Dl-3-n-butylphthalide (NBP) has anti-inflammatory and antioxidative properties. However, whether NBP can protect skeletal muscles from limb I/R injury and the mechanism in mediating the action of NBP treatment still remain to be investigated, which are the focuses of the current study. Methods: The model of limb I/R injury was established and H&E staining was adopted to assess the pathological changes in skeletal muscles following limb I/R injury. Additionally, the W/D ratio of muscle tissue was also measured. ELISA and biochemical tests were carried out to measure the levels of inflammatory cytokines and oxidative stress in mouse models of limb I/R injury. Moreover, the levels of the HMGB1/TLR4/NF-κB pathway-related proteins were also determined using immunohistochemistry and immunoblotting. Results: It was established that NBP treatment alleviated I/R-induced pathological changes in muscular tissue of mice, accompanied by lower W/D ratio of skeletal muscular tissue. Meanwhile, the limb I/R-induced inflammation and oxidative stress in skeletal muscles of mice were also inhibited by NBP. Mechanistic study indicated that the alleviatory effect of NBP was ascribed to inactivation of the HMGB1/TLR4/NF-κB pathway. Conclusions: Our findings highlighted the potential of NBP as a novel strategy for limb I/R-driven muscle tissue damages by suppressing inflammatory response and oxidative stress via the HMGB1/TLR4/NF-κB pathway.

Knockdown of lncRNA HOXA-AS3 Suppresses the Progression of Atherosclerosis via Sponging miR-455-5p.

BACKGROUND: Atherosclerosis can lead to multiple cardiovascular diseases, especially myocardial infarction. Long noncoding RNAs (lncRNAs) are involved in multiple diseases, including atherosclerosis. LncRNA HOXA-AS3 was found to be notably upregulated in atherosclerosis. However, the biological function of HOXA-AS3 during the occurrence and development of atherosclerosis remains unclear. MATERIALS AND METHODS: Human vascular endothelial cells (HUVECs) were treated with oxidized low-density lipoprotein (oxLDL) to mimic atherosclerosis in vitro. Gene and protein expressions in HUVECs were detected by RT-qPCR and Western blot, respectively. Cell proliferation was tested by CCK-8 and Ki67 staining. Cell apoptosis and cycle were measured by flow cytometry. Additionally, the correlation between HOXA-AS3 and miR-455-5p was confirmed by dual luciferase report assay and RNA pull-down. Finally, in vivo model of atherosclerosis was established to confirm the function of HOXA-AS3 during the development of atherosclerosis in vivo. RESULTS: LncRNA HOXA-AS3 was upregulated in oxLDL-treated HUVECs. In addition, oxLDL-induced growth inhibition of HUVECs was significantly reversed by knockdown of HOXA-AS3. Consistently, oxLDL notably induced G1 arrest in HUVECs, while this phenomenon was greatly reversed by HOXA-AS3 siRNA. Furthermore, downregulation of HOXA-AS3 notably inhibited the progression of atherosclerosis through mediation of miR-455-5p/p27 Kip1 axis. Besides, silencing of HOXA-AS3 notably relieved the symptom of atherosclerosis in vivo. CONCLUSION: Downregulation of HOXA-AS3 significantly suppressed the progression of atherosclerosis via regulating miR-455-5p/p27 Kip1 axis. Thus, HOXA-AS3 might serve as a potential target for the treatment of atherosclerosis.

Carboxypeptidase A4 promotes cardiomyocyte hypertrophy through activating PI3K-AKT-mTOR signaling.

Carboxypeptidase A4 (CPA4) is a member of the metallocarboxypeptidase family. Current studies have identified the roles of CPA4 in cancer biology and insulin sensitivity. However, the roles of CPA4 in other diseases are not known. In the present study, we investigated the roles of CPA4 in cardiac hypertrophy. The expression of CPA4 was significantly increased in the hypertrophic heart tissues of human patients and isoproterenol (ISO)-induced hypertrophic heart tissues of mice. We next knocked down Cpa4 with shRNA or overexpressed Cpa4 using adenovirus in neonatal rat cardiomyocytes and induced cardiomyocyte hypertrophy with ISO. We observed that Cpa4 overexpression promoted whereas Cpa4 knockdown reduced ISO-induced growth of cardiomyocyte size and overexpression of hypertrophy marker genes, such as myosin heavy chain ß (ß-Mhc), atrial natriuretic peptide (Anp), and brain natriuretic peptide (Bnp). Our further mechanism study revealed that the mammalian target of rapamycin (mTOR) signaling was activated by Cpa4 in cardiomyocytes, which depended on the phosphoinositide 3-kinase (PI3K)-AKT signaling. Besides, we showed that the PI3K-AKT-mTOR signaling was critically involved in the roles of Cpa4 during cardiomyocyte hypertrophy. Collectively, these results demonstrated that CPA4 is a regulator of cardiac hypertrophy by activating the PI3K-AKT-mTOR signaling, and CPA4 may serve as a promising target for the treatment of hypertrophic cardiac diseases.

HTR2A promotes the development of cardiac hypertrophy by activating PI3K-PDK1-AKT-mTOR signaling.

5-Hydroxytryptamine receptor 2A (HTR2A) is a central regulator of fetal brain development and cognitive function in adults. However, the roles of HTR2A in the cardiovascular system are not fully understood. Here in this study, we explored the function of HTR2A in cardiac hypertrophy. Significantly, the expression levels of HTR2A mRNA and protein levels were upregulated in hypertrophic hearts of human patients. Besides, the expression of HTR2A was also upregulated in isoproterenol (ISO)-induced cardiac hypertrophy in the mouse. Next, the expression of HTR2A was knocked down with shRNA or overexpressed with adenovirus in neonatal rat cardiomyocytes, and ISO was used to induce cardiomyocyte hypertrophy. We showed that HTR2A knockdown repressed ISO-induced cardiomyocyte hypertrophy, which was demonstrated by decreased cardiomyocyte size and repressed expression of hypertrophic fetal genes (e.g., myosin heavy chain beta (ß-Mhc), atrial natriuretic peptide (Anp), and brain natriuretic peptide (Bnp)). By contrast, HTR2A overexpression promoted cardiomyocyte hypertrophy. Of note, we observed that HTR2A promoted the activation (phosphorylation) of AKT-mTOR (mammalian target of rapamycin) signaling in cardiomyocytes, and repression of AKT-mTOR with perifosine or rapamycin blocked the effects of HTR2A on cardiomyocyte hypertrophy. Finally, we showed that HTR2A regulated AKT-mTOR signaling through activating the PI3K-PDK1 pathway, and inhibition of either PI3K or PDK1 blocked the roles of HTR2A in regulating AKT-mTOR signaling and cardiomyocyte hypertrophy. Altogether, these findings demonstrated that HTR2A activated PI3K-PDK1-AKT-mTOR signaling and promoted cardiac hypertrophy.

Smooth muscle SIRT1 reprograms endothelial cells to suppress angiogenesis after ischemia.

Objective: Vascular smooth muscle cells (VSMCs) undergo the phenotypic changes from contractile to synthetic state during vascular remodeling after ischemia. SIRT1 protects against stress-induced vascular remodeling via maintaining VSMC differentiated phenotype. However, the effect of smooth muscle SIRT1 on the functions of endothelial cells (ECs) has not been well clarified. Here, we explored the role of smooth muscle SIRT1 in endothelial angiogenesis after ischemia and the underlying mechanisms. Methods: We performed a femoral artery ligation model using VSMC specific human SIRT1 transgenic (SIRT1-Tg) and knockout (KO) mice. Angiogenesis was assessed in in vivo by quantification of the total number of capillaries, wound healing and matrigel plug assays, and in vitro ECs by tube formation, proliferation and migration assays. The interaction of HIF1α with circRNA was examined by using RNA immunoprecipitation, RNA pull-down and in situ hybridization assays. Results: The blood flow recovery was significantly attenuated in SIRT1-Tg mice, and markedly improved in SIRT1-Tg mice treated with SIRT1 inhibitor EX527 and in SIRT1-KO mice. The density of capillaries significantly decreased in the ischemic gastrocnemius of SIRT1-Tg mice compared with SIRT1-KO and WT mice, with reduced expression of VEGFA, which resulted in decreased number of arterioles. We identified that the phenotypic switching of SIRT1-Tg VSMCs was attenuated in response to hypoxia, with high levels of contractile proteins and reduced expression of the synthetic markers and NG2, compared with SIRT1-KO and WT VSMCs. Mechanistically, SIRT1-Tg VSMCs inhibited endothelial angiogenic activity induced by hypoxia via the exosome cZFP609. The cZFP609 was delivered into ECs, and detained HIF1α in the cytoplasm via its interaction with HIF1α, thereby inhibiting VEGFA expression and endothelial angiogenic functions. Meantime, the high cZFP609 expression was observed in the plasma of the patients with atherosclerotic or diabetic lower extremity peripheral artery disease, associated with reduced ankle-brachial index. Knockdown of cZFP609 improved blood flow recovery after hindlimb ischemia in SIRT1-Tg mice. Conclusions: Our findings demonstrate that SIRT1 may impair the plasticity of VSMCs. cZFP609 mediates VSMCs to reprogram endothelial functions, and serves as a valuable indicator to assess the prognosis and clinical outcomes of ischemic diseases.

Comparison of false negative rates between 100% rapid review and 10% random full rescreening as internal quality control methods in cervical cytology screening.

OBJECTIVE: To compare and contrast the 100% partial rapid review (RR) method against 10% randomly selected full rescreening to determine the better means of internal quality control (QC) by which the cervical screening laboratory should adopt. STUDY DESIGN: A total of 4,150 conventional Pap smears reported as negative during 2003 were 100% rescreened rapidly in large field jumps. From this cohort, 415 (10%) were selected again randomly for full rescreening. The false negative rates (FNRs) were calculated and compared accordingly. RESULTS: The 100% RR method reclassified 30 false negative (FN) cases, while the 10% random full rescreening reclassified only 5. The FNRs were calculated to be 8.2% and 12.9%, respectively. A chi2 test (p < 0.05) showed significant difference between the FNRs calculated by both methods. CONCLUSION: The 100% RR proved to be a better method for QC purpose in reducing the FNRs and detected more missing cases. This study is the first of its type from Hong Kong and confirms previous publications from elsewhere that 100% RR is a better method for internal QC in cervical cytology screening.