Melastoma dodecandrum lour. Protects against cerebral ischemia-reperfusion injury by ameliorating oxidative stress and endoplasmic reticulum stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Melastoma dodecandrum Lour. (MD), a traditional Chinese medicine used by the She ethnic group, has been used to treat cerebral ischemia-reperfusion (CIR) injury due to its efficacy in promoting blood circulation and removing blood stasiss; however, the therapeutic effects and mechanisms of MD in treating CIR injury remain unclear. AIM: To investigate the protective effects of MD on CIR injury, in addition to its impact on oxidative stress, endoplasmic reticulum (ER) stress, and cell apoptosis. MATERIALS AND METHODS: The research was conducted using both cell experiments and animal experiments. The CCK-8 method, immunofluorescence staining, and flow cytometry were used to analyze the effects of MD-containing serum on oxygen-glucose deprivation/reperfusion (OGD/R)-induced PC12 cell viability, reactive oxygen species (ROS) clearance, anti-inflammatory, neuroprotection and inhibition of apoptosis. Furthermore, 2,3,5-Triphenyl tetrazolium chloride staining, hematoxylin and eosin staining, Nissl staining, and immunohistochemistry were used to detect infarct size, pathological changes, Nissl corpuscula and neuronal protein expression in middle cerebral artery occlusion (MCAO) rats. Polymerase chain reaction and Western Blotting were conducted in cell and animal experiments to detect the expression levels of ER stress-related genes and proteins. RESULTS: The MD extract enhanced the viability of PC12 cells under OGD/R modeling, reduced ROS and IL-6 levels, increased MBP levels, and inhibited cell apoptosis. Furthermore, MD improved the infarct area in MCAO rats, increased the number of Nissl bodies, and regulated neuronal protein levels including Microtubule-Associated Protein 2 (MAP-2), Myelin Basic Protein (MBP), Glial Fibrillary Acidic Protein (GFAP), and Neurofilament 200 (NF200). Additionally, MD could regulate the expression levels of oxidative stress proteins malondialdehyde (MDA), nitric oxide (NO), superoxide dismutase (SOD), and catalase (CAT). Both cell and animal experiments demonstrated that MD could inhibit ER stress-related proteins (GRP78, ATF4, ATF6, CHOP) and reduce cell apoptosis. CONCLUSION: This study confirmed that the therapeutic mechanism of the MD extract on CIR injury was via the inhibition of oxidative stress and the ER stress pathway, in addition to the inhibition of apoptosis.

Naoxintong capsule accelerates mitophagy in cerebral ischemia-reperfusion injury via TP53/PINK1/PRKN pathway based on network pharmacology analysis and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: The incidence and mortality of cerebrovascular diseases are increasing year by year. Cerebral ischemia-reperfusion injury (CIRI) is common in patients with ischemic stroke. Naoxintong (NXT) is composed of a variety of Chinese medicines and has the ability to treat CIRI. AIM OF THE STUDY: The aim of this study is to investigate whether NXT regulates mitophagy in CIRI based on network pharmacology analysis and experimental validation. MATERIALS AND METHODS: Oxygen and glucose deprivation/re-oxygenation (OGD/R, 2/22 h) model of PC12 cells and transient middle cerebral artery occlusion (tMCAO, 2/22 h) model of rats were established. Pharmacodynamic indicators include neurological deficit score, 2,3,5-triphenyte-trazoliumchloride (TTC) staining, hematoxylin-eosin (HE) staining and cell viability. Network pharmacology was used to predict pharmacological mechanisms. Pharmacological mechanism indexes include transmission electron microscopy (TEM), drug affinity responsive target stability (DARTS), cellular thermal shift assay (CETSA), immunohistochemistry (IHC), western blot (WB) and immunofluorescence (IF). Kevetrin (an agonists of p53) and pifithrin-α (an inhibitor of p53) used to detect the key role of p53 in mitophagy of NXT. RESULTS: NXT (1% serum containing NXT and 110 mg/kg) improved the damage of OGD/R PC12 cells and tMCAO rats, and this protective effect was related to the anti-oxidation and ability to promote mitophagy of NXT. NXT and pifithrin-α increased the expression of promoting-mitophagy targets (PINK1, PRKN and LC3B) and inhibited the expression of inhibiting-mitophagy targets (p52) via restraining p53, and finally accelerated mitophagy caused by CIRI. CONCLUSION: This study demonstrates that NXT promotes mitophagy in CIRI through restraining p53 and promoting PINK1/PRKN in vivo and in vitro.

[Buyang Huanwu Decoction attenuates cerebral ischemia-reperfusion injury in rats via miR-26a-5p mediated PTEN/PI3K/Akt signaling pathway].

This study aims to investigate the mechanism of Buyang Huanwu Decoction in treatment of cerebral ischemia-reperfusion injury in rats. A total of 180 SD rats were randomly divided into 5 different groups: sham group, model group, Buyang Huanwu Decoction group, Buyang Huanwu Decoction + miR-26a-5p agomir(agomir) group, Buyang Huanwu Decoction + miR-26a-5p agomir negative control(agomir NC) group. There were 36 rats in each group. Each group was then subdivided into three subgroups for the duration of reperfusion(3, 7, 14 d). A ligature-induced middle cerebral artery occlusion(MCAO) model was carried out on all groups other than sham group. Reperfusion was performed following ischemia for 90 min. Buyang Huanwu Decoction group, agomir group, and agomir NC group were given Buyang Huanwu Decoction twice daily by gavage 24 h after the formation of the model. Sham group and model group were given an equal amount of physiological saline by gavage until the day before sacrifice. At 24 h after ischemia induction, miR-26a-5p agomir was injected into the lateral ventricle in agomir group, miR-26a-5p NC in agomir NC group, and equal amounts of physiological saline in the other groups. 24 h after ischemia induction, BrdU was intraperitoneally injected once daily until the day before sacrifice. Modified neurological severity score(mNSS) was used to evaluate neurological deficits, 2,3,5-triphenyltetrazolium chloride(TTC) staining was used to determine the cerebral infarct volume, TUNEL staining was used to assess the apoptosis of parenchymal ischemic brain tissue, and double immunofluorescence staining was used to examine BrdU/NeuN double positive neurons in the parenchymal ischemic brain tissue to evaluate the neuronal regeneration. We employed a luciferase reporter assay to identify and validate that the target gene of miR-26a-5p is PTEN. Real-time quantitative polymerase chain reaction(RT-qPCR) was used to assess gene expression levels of PTEN and miR-26a-5p and Western blot to assess the protein levels of PTEN, PI3K, p-PI3K, Akt, and p-Akt. The results revealed that compared with model group, Buyang Huanwu Decoction treatment promoted neural function recovery, reduced the cerebral infarct volume, increased the number of BrdU~+/NeuN~+ neurons, upregulated the expression of miR-26a-5p, regulated the PTEN/PI3K/Akt signaling pathway, and promoted neuronal regeneration in the cerebral ischemia-reperfusion rats. These effects were significantly enhanced after lateral ventricle injection of miR-26a-5p agomir. The findings prove that Buyang Huanwu Decoction treatment can promote neural function recovery, reduce the cerebral infarct volume, and promote neuronal regeneration in a cerebral ischemia-reperfusion rat model, which is likely to be achieved via miR-26a-5p mediated PTEN/PI3K/Akt signaling pathway.

Autophagy in myocardial ischemia and ischemia/reperfusion.

Myocardial infarction (MI) is a life-threatening condition that leads to loss of viable heart tissue. The best way to treat acute MI and limit the infarct size is to re-open the occluded coronary artery and restore the supply of oxygenated and nutrient-rich blood, but reperfusion can cause additional damage. Autophagy is an intracellular process that recycles damaged cytoplasmic components (molecules and organelles) by loading them into autophagosomes and degrading them in autolysosomes. Autophagy is increased in in vivo animal models of permanent ischemia and ischemia/reperfusion but by different molecular mechanisms. While autophagy is protective during permanent ischemia, it is detrimental during ischemia/reperfusion. Its modulation is being investigated as a potential target to reduce reperfusion injury. This review provides a synopsis of the current knowledge about autophagy, summarizes findings specifically in permanent ischemia and ischemia/reperfusion, and briefly discusses the potential implication of experimental findings.

Combined effect of intermittent hemostasis and a modified external hemorrhage control device in a lethal swine model.

Background: Non-compressible torso hemorrhage (NCTH) presents the ultimate challenge in pre-hospital care. While external hemorrhage control devices (EHCDs) such as the Abdominal Aortic and Junctional Tourniquet (AAJT) and SAM Junctional Tourniquet (SJT) have been invented, the current design and application strategy requires further improvement. Therefore, researchers devised a novel apparatus named Modified EHCD (M-EHCD) and implemented intermittent hemostasis (IH) as a preventive measure against ischemia-reperfusion injury. The objective of this study was to ascertain the combined effect of M-EHCD and IH on the hemostatic effect of NCTH. Methods: Eighteen swine were randomized to M-EHCD, AAJT or SJT. The NCTH model was established by inducing Class Ⅲ hemorrhagic shock and performing a hemi-transection of common femoral artery (CFA). EHCDs were rapidly fastened since the onset of free bleeding (T0min). The IH strategy was implemented by fully releasing M-EHCD at T40min, T70min and T100min, respectively, whereas AAJT and SJT maintained continuous hemostasis (CH) until T120min. All groups underwent CFA bridging at T110min, and EHCDs were removed at T120min. Reperfusion lasted for 60 min, after which euthanasia was performed. Hemodynamics, intra-vesical pressure (IVP), and blood samples were collected periodically. Histological examinations were also conducted. Results: M-EHCD demonstrated the fastest application time (M-EHCD: 26.38 ± 6.32s vs. SJT: 30.84 ± 5.62s vs. AAJT: 54.28 ± 5.45s, P < 0.001) and reduced free blood loss (M-EHCD: 17.77 ± 9.85g vs. SJT: 51.80 ± 33.70g vs. AAJT: 115.20 ± 61.36g, P = 0.011) compared to SJT and AAJT. M-EHCD exhibited inhibitory effects on heart rate (M-EHCD: 91.83 ± 31.61bpm vs. AAJT: 129.00 ± 32.32bpm vs. SJT: 135.17 ± 21.24bpm, P = 0.041) and shock index. The device's external pressure was lowest in M-EHCD and highest in SJT (P = 0.001). The resultant increase in IVP were still the lowest in M-EHCD (M-EHCD: -0.07 ± 0.45 mmHg vs. AAJT: 27.04 ± 5.03 mmHg vs. SJT: 5.58 ± 2.55 mmHg, P < 0.001). Furthermore, M-EHCD caused the least colonic injury (M-EHCD: 1.17 ± 0.41 vs. AAJT: 2.17 ± 0.41 vs. SJT: 2.17 ± 0.41, P = 0.001). The removal of M-EHCD showed the slightest impact on pH (P < 0.001), while AAJT group was more susceptible to the lethal triad based on the arterial lactate and thrombelastogram results. Conclusions: M-EHCD + IH protected the organs and reduced the risk of the lethal triad by decreasing disruptions to IVP, hemodynamics, acid-base equilibrium and coagulation. M-EHCD + IH was superior to the hemostatic safety and efficacy of AAJT/SJT + CH.

Retinal ischemia-reperfusion injury and pretreatment with Lycium barbarum glycopeptide.

AIM: To investigate the antioxidant protective effect of Lycium barbarum glycopeptide (LbGP) pretreatment on retinal ischemia-reperfusion (I/R) injury (RIRI) in rats. METHODS: RIRI was induced in Sprague Dawley rats through anterior chamber perfusion, and pretreatment involved administering LbGP via gavage for 7d. After 24h of reperfusion, serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and creatinine (CREA) levels, retinal structure, expression of Caspase-3 and Caspase-8, superoxide dismutase (SOD) activity, and malondialdehyde (MDA) in the retina were measured. RESULTS: The pretreatment with LbGP effectively protected the retina and retinal tissue from edema and inflammation in the ganglion cell layer (GCL) and nerve fiber layer (NFL) of rats subjected to RIRI, as shown by light microscopy and optical coherence tomography (OCT). Serum AST was higher in the model group than in the blank group (P=0.042), but no difference was found in ALT, AST, and CREA across the LbGP groups and model group. Caspase-3 expression was higher in the model group than in the blank group (P=0.006), but no difference was found among LbGP groups and the model group. Caspase-8 expression was higher in the model group than in the blank group (P=0.000), and lower in the 400 mg/kg LbGP group than in the model group (P=0.016). SOD activity was lower in the model group than in the blank group (P=0.001), and the decrease was slower in the 400 mg/kg LbGP group than in the model group (P=0.003). MDA content was higher in the model group than in the blank group (P=0.001), and lower in the 400 mg/kg LbGP group than in the model group (P=0.016). The pretreatment with LbGP did not result in any observed liver or renal toxicity in the model. CONCLUSION: LbGP pretreatment exhibits dose-dependent anti-inflammatory, and antioxidative effects by reducing Caspase-8 expression, preventing declines of SOD activity, and decreasing MDA content in the RIRI rat model.

Transcriptomic signatures during normothermic liver machine perfusion correspond with graft quality and predict the early graft function.

BACKGROUND: A better understanding of the molecular events during liver normothermic machine perfusion (NMP) is warranted to develop a data-based approach for the identification of biomarkers representative of graft quality and posttransplant outcome. We analysed the dynamic transcriptional changes during NMP and linked them to clinical and biochemical parameters. METHODS: 50 livers subjected to NMP for up to 24 h were enrolled. Bulk RNA sequencing was performed in serial biopsies collected pre and during NMP, and after reperfusion. Perfusate was sampled to monitor liver function. qPCR and immunohistochemistry were performed to validate findings. Molecular profiles were compared between transplanted and non-transplanted livers, and livers with and without early allograft dysfunction. FINDINGS: Pathways related to immune and cell stress responses, cell trafficking and cell regulation were activated during NMP, while cellular metabolism was downregulated over time. Anti-inflammatory responses and genes involved in tissue remodelling were induced at later time-points, suggesting a counter-response to the immediate damage. NMP strongly induced a gene signature associated with ischemia-reperfusion injury. A 7-gene signature corresponds with the benchmarking criteria for transplantation or discard at 6 h NMP (area under curve 0.99). CD274 gene expression (encoding programmed cell-death ligand-1) showed the highest predictive value. LEAP2 gene expression at 6 h NMP correlated with impaired graft function. INTERPRETATION: Assessment of gene expression markers could serve as a reliable tool to evaluate liver quality during NMP and predicts early graft function after transplantation. FUNDING: The research was supported by "In Memoriam Dr. Gabriel Salzner Stiftung", Tiroler Wissenschaftsfond, Jubiläumsfonds-Österreichische Nationalbank and MUI Start grant.

Time varying characteristic in somatosensory evoked potentials as a biomarker of spinal cord ischemic-reperfusion injury in rat.

Spinal cord ischemic-reperfusion injury (SCIRI) could occurs during surgical procedures without detection, presenting a complex course and an unfavorable prognosis. This may lead to postoperative sensory or motor dysfunction in areas innervated by the spinal cord, and in some cases, permanent paralysis. Timely detection of SCIRI and immediate waring can help surgeons implement remedial intervention to prevent irreversible spinal cord injury. Therefore, it is crucial to develop a precise and effective method for early detection of SCIRI. This study utilized rat models to simulate intraoperative SCIRI and employed somatosensory evoked potentials (SEP) for continuous monitoring during surgery. In this study, SEP signal changes were examined in six groups with varying severities of SCIRI and one normal control group. SEP signal changes were examined during operations in different groups and correlated with postoperative behavioral and histopathological data. The result demonstrated specific changes in SEP signals during SCIRI, termed as time-varying characteristics, which are associated with the duration of ischemia and subsequent reperfusion. Time-varying characteristics in SEP could potentially serve as a new biomarker for the intraoperative detection of SCIRI. This finding is significant for clinical surgeons to identify and guide early intervention of SCIRI timely. Additionally, this measurement is easily translatable to clinical application.

Canine mesenchymal stem cell-derived exosomes attenuate renal ischemia-reperfusion injury through miR-146a-regulated macrophage polarization.

Introduction: The most common factor leading to renal failure or death is renal IR (ischemia-reperfusion). Studies have shown that mesenchymal stem cells (MSCs) and their exosomes have potential therapeutic effects for IR injury by inhibiting M1 macrophage polarization and inflammation. In this study, the protective effect and anti-inflammatory mechanism of adipose-derived mesenchymal stem cell-derived exosomes (ADMSC-Exos) after renal IR were investigated. Method: Initially, ADMSC-Exos were intravenously injected into IR experimental beagles, and the subsequent assessment focused on inflammatory damage and macrophage phenotype. Furthermore, an in vitro inflammatory model was established by inducing DH82 cells with LPS. The impact on inflammation and macrophage phenotype was then evaluated using ADMSC and regulatory miR-146a. Results: Following the administration of ADMSC-Exos in IR canines, a shift from M1 to M2 macrophage polarization was observed. Similarly, in vitro experiments demonstrated that ADMSC-Exos enhanced the transformation of LPS-induced macrophages from M1 to M2 type. Notably, the promotion of macrophage polarization by ADMSC-Exos was found to be attenuated upon the inhibition of miR-146a in ADMSC-Exos. Conclusion: These findings suggest that miR-146a plays a significant role in facilitating the transition of LPS-induced macrophages from M1 to M2 phenotype. As a result, the modulation of macrophage polarization by ADMSC-Exos is achieved via the encapsulation and conveyance of miR-146a, leading to diminished infiltration of inflammatory cells in renal tissue and mitigation of the inflammatory reaction following canine renal IR.

Neutrophil extracellular traps and citrullinated fibrinogen contribute to injury in a porcine model of limb ischemia and reperfusion.

Background: Ischemia/reperfusion injury (IRI) is a complex pathological process, triggered by the restoration of blood flow following an interrupted blood supply. While restoring the blood flow is the only option to salvage the ischemic tissue, reperfusion after a prolonged period of ischemia initiates IRI, triggering a cascade of inflammatory responses ultimately leading to neutrophil recruitment to the inflamed tissue, where they release neutrophil extracellular traps (NETs). NETs are web-like structures of decondensed chromatin and neutrophilic proteins, including peptidyl-arginine deiminase 2 and 4 (PAD2, PAD4), that, once outside, can citrullinate plasma proteins, irreversibly changing their conformation and potentially their function. While the involvement of NETs in IRI is known mainly from rodent models, we aimed to determine the effect of NET formation and especially PADs-mediated extracellular protein citrullination in a porcine model of limb IRI. Methods: We conducted our study on amputated pig forelimbs exposed to 1 h or 9 h of ischemia and then reperfused in vivo for 12 h. Limb weight, edema formation, compartmental pressure were measured, and skeletal muscle was analyzed by immunofluorescence (TUNEL assay and dystrophin staining) to evaluate tissue damage. Fibrin tissue deposition, complement deposition and NETs were investigated by immunofluorescence. Citrullinated plasma proteins were immunoprecipitated and citrullinated fibrinogen was identified in the plasma by Western blot and in the tissue by immunofluorescence and Western blot. Results: Our data consolidate the involvement of NETs in a porcine model of limb IRI, correlating their contribution to damage extension with the duration of the ischemic time. We found a massive infiltration of NETs in the group subjected to 9 h ischemia compared to the 1 h and citrullinated fibrinogen levels, in plasma and tissue, were higher in 9 h ischemia group. We propose fibrinogen citrullination as one of the mechanisms contributing to the worsening of IRI. NETs and protein citrullination represent a potential therapeutic target, but approaches are still a matter of debate. Here we introduce the idea of therapeutic approaches against citrullination to specifically inhibit PADs extracellularly, avoiding the downstream effects of hypercitrullination and keeping PADs' and NETs' intracellular regulatory functions.

Cardiac-targeted and ROS-responsive liposomes containing puerarin for attenuating myocardial ischemia-reperfusion injury.

Aim: This study aimed to construct an ischemic cardiomyocyte-targeted and ROS-responsive drug release system to reduce myocardial ischemia-reperfusion injury (MI/RI).Methods: We constructed thioketal (TK) and cardiac homing peptide (CHP) dual-modified liposomes loaded with puerarin (PUE@TK/CHP-L), which were expected to deliver drugs precisely into ischemic cardiomyocytes and release drugs in response to the presence of high intracellular ROS levels. The advantages of PUE@TK/CHP-L were assessed by cellular pharmacodynamics, in vivo fluorescence imaging and animal pharmacodynamics.Results: PUE@TK/CHP-L significantly inhibited apoptosis and ferroptosis in H/R-injured cardiomyocytes and also actively targeted ischemic myocardium. Based on these advantages, PUE@TK/CHP-L could significantly enhance the drug's ability to attenuate MI/RI.Conclusion: PUE@TK/CHP-L had potential clinical value in the precise treatment of MI/RI.

[Box: see text].

Myrtenol-Loaded Fatty Acid Nanocarriers Protect Rat Brains Against Ischemia-Reperfusion Injury: Antioxidant and Anti-Inflammatory Effects.

This research investigated the preventive effects of myrtenol (MYR), fatty acid nanocarriers (FANC), and myrtenol-loaded FANC (MYR + FANC) on neurological disturbance, stroke volume, the levels of malondialdehyde (MDA), superoxide dismutase (SOD), and tumor necrosis factor-alpha (TNF-α) in the brain with ischemia-reperfusion injuries induced by middle cerebral artery occlusion (MCAO) in rats. Seventy two Wistar male rats were divided into six main groups. The groups were sham, ischemia-reperfusion group (MACO), MACO-MYR (50 mg/kg), MACO-FANC (50 and 100 mg/kg), and MACO-MYR + FANC (50 mg/kg). Stroke volume, neurological deficit scores, and the brain levels of MDA, SOD, and TNF-α were examined with TTC staining, observation, and ELISA, respectively. Pretreatment with MYR, FANC (100 mg/kg), and MYR + FANC reduced the neurological deficit score and cerebral infarction volume. MYR, FANC (100 mg/kg), and MYR + FANC pretreatment increased and decreased brain SOD and MDA levels compared to MACO group, respectively. The TNF-α level decreased in the MYR + FANC group compared to MCAO and MCAO-MYR groups in the brain. The use of FANC (100 mg/kg), MYR, and MYR + FANC has protective effects against oxidative stress and ischemia-reperfusion injury. FANC probably improve the bioavailability of MYR, as MYR+ FANC had more therapeutic effects on the reduction of ischemia-reperfusion injuries, inflammation, and oxidative stress.

High­fat diet­induced LCN2 exacerbates myocardial ischemia­reperfusion injury by enhancing platelet activation.

Following acute myocardial infarction, the recovery of blood flow leads to myocardial ischemia­reperfusion (MI/R) injury, which is primarily characterized by the activation of inflammatory signals, microvascular obstruction, increased oxidative stress and excessive Ca2+ overload. It has also been demonstrated that platelets can exacerbate MI/R injury by releasing reactive oxygen species, inflammatory factors and chemokines, while also obstructing microvessels through thrombus formation. As a bioactive molecule with proinflammatory and chemotactic properties, lipocalin 2 (LCN2) exhibits a positive correlation with obesity, hyperglycemia, hypertriglyceridemia and insulin resistance index, which are all significant risk factors for ischemic cardiomyopathy. Notably, the potential role of LCN2 in promoting atherosclerosis may be related to its influence on the function of macrophages, smooth muscle cells and endothelial cells, but its effect on platelet function has not yet been reported. In the present study, the effect of a high­fat diet (HFD) on LCN2 expression was determined by detecting LCN2 expression levels in the liver and serum samples of mice through reverse transcription­quantitative PCR and enzyme linked immunosorbent assay, respectively. The effect of LCN2 on platelet function was evaluated by examining whether LCN2 affected platelet activation, aggregation, adhesion, clot retraction and P­selectin expression. To determine whether LCN2 aggravated MI/R injury in HFD­fed mice by affecting platelet and inflammatory cell recruitment, wild­type and LCN2 knockout mice fed a HFD were subjected to MI/R injury, then hearts were collected for hematoxylin and eosin staining and 2,3,5­triphenyltetrazolium chloride staining, and immunohistochemistry was employed to detect the expression of CD42b, Ly6G, CD3 and B220. Based on observing the upregulation of LCN2 expression in mice fed a HFD, the present study further confirmed that LCN2 could accelerate platelet activation, aggregation and adhesion. Moreover, in vivo studies validated that knockout of LCN2 not only mitigated MI/R injury, but also inhibited the recruitment of platelets and inflammatory cells in myocardial tissue following ischemia­reperfusion. In conclusion, the current findings suggested that the effect of HFD­induced LCN2 on aggravating MI/R injury may totally or partially dependent on its promotion of platelet function.

A novel framework for uncovering the coordinative spectrum-effect correlation of the effective components of Yangyin Tongnao Granules on cerebral ischemia-reperfusion injury in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ischemic stroke is currently a major public health hazard.Yangyin Tongnao Granules (YYTN), a traditional Chinese medicinal prescription, exerts potential therapeutic effects on subsequent cerebral ischemia-reperfusion injury (CIRI) after ischemic stroke. However, further studies are required to comprehend the underlying mechanism of YYTN for treating CIRI and the associated spectrum-effect mechanisms. AIM OF THE STUDY: To investigate the coordinated correlation between the fingerprint and the pharmacodynamic indexes of the effective components (total flavonoids, total saponins, total alkaloids, and total phenolic acids) in YYTN for treating CIRI in rats. METHODS: The fingerprints of five specific components (ligustrazine, puerarin, ferulic acid, calycosin, and formononetin) of YYTN in rats with middle cerebral artery occlusion (MCAO) were established using high-performance liquid chromatography (HPLC), and their peak areas were quantified in plasma samples. The pharmacodynamic indexes of tumor necrosis factor-alpha (TNF-α), cytochrome c (Cyt-C), and total superoxide dismutase (T-SOD) were integrated using the Criteria Importance Through Intercriteria Correlation (CRITIC) method to create a comprehensive evaluation index. Spectrum-effect correlation was analyzed by performing gray relation analysis (GRA), correlation analysis (CA), and partial least squares regression (PLSR). The Borda method was then applied to integrate the obtained results. RESULTS: In MCAO rats, the effective components of YYTN reduced TNF-α and Cyt-C and increased T-SOD, which indicates their anti-inflammatory, antiapoptotic, and antioxidant effects. Spectrum-effect CA revealed certain associations between the chromatographic peaks of the five main components and the comprehensive pharmacodynamic evaluation index. Of these components, formononetin displayed the highest correlation, whereas ferulic acid exhibited the lowest correlation. All components showed a positive correlation. Using the Borda method, the components were ranked as follows based on correlation: formononetin > calycosin > ligustrazine > puerarin > ferulic acid. CONCLUSIONS: The effective components of YYTN exhibited synergistic effects in the treatment of MCAO rats, which could potentially be attributed to their multitarget and multipathway mechanisms. The Borda method-based spectrum-effect correlation analysis provides a coordinated approach to investigate the relationship between fingerprint and pharmacodynamics of traditional Chinese medicine (TCM).

Borneol promotes berberine-induced cardioprotection in a rat model of myocardial ischemia/reperfusion injury via inhibiting P-glycoprotein expression.

Berberine is reported to protect the heart against ischemia/reperfusion (I/R) injury, although efficacy is limited by low bioavailability. This study aims to determine whether borneol, a classic guiding drug, can enhance the cardioprotection induced by berberine and to clarify the underlying mechanisms involving P-glycoprotein (P-gp) in the heart. Adult male Sprague Dawley rats were gavaged with berberine (200 mg/kg) with or without borneol (100 mg/kg) for 7 consecutive days. A rat model of myocardial I/R injury was established by 30 min left coronary artery occlusion followed with 120 min reperfusion. The arrhythmia score, cardiac enzyme content, and myocardial infarct size were determined following reperfusion. Heart tissues were collected for western blot and immunofluorescence analyses to measure the protein expression levels of Bcl-2, Bax, and P-gp. The results showed that administration of berberine protected the heart against I/R injury, as demonstrated by lower arrhythmia scores, serum cTnI contents, myocardial infarct size, and cardiomyocytes apoptosis. Moreover, borneol substantially enhanced the cardioprotective effects of berberine. Western blot and immunofluorescence analyses showed that both berberine and I/R injury did not alter P-gp expression in heart. In contrast, borneol combined with berberine significantly reduced P-gp levels by 43.4% (P = 0.0240). Interestingly, treatment with borneol alone decreased P-gp levels, but did not protect against myocardial I/R injury. These findings suggest that borneol, as an adjuvant drug, improved the cardioprotective effects of berberine by inhibiting P-gp expression in heart. Borneol combined with berberine administration provides a new strategy to protect the heart against I/R injury.

Bone marrow mesenchymal stem cells with PTBP1 knockdown protect against cerebral ischemia-reperfusion injury by inhibiting ferroptosis via the JNK/P38 pathway in rats.

Over the years, the neuroprotective potential of bone marrow mesenchymal stem cells (BMSCs) in acute ischemic stroke has attracted significant attention. However, BMSCs face challenges like short metabolic cycles and low survival rates post-transplant. Polypyrimidine tract-binding protein 1 (PTBP1) is an immunomodulatory RNA-binding protein that regulates the cell cycle and increases cell viability. This study investigated the protective effects and underlying mechanism of PTBP1 knockdown in BMSCs (PTBP1KD-BMSCs) following ischemia-reperfusion injury (IRI) in neurons. BMSCs were isolated from Sprague-Dawley rat femurs and characterized through flow cytometry and differentiation induction. PTBP1 knockdown inhibited BMSCs proliferation. Co-culture with PTBP1KD-BMSCs decreased reactive oxygen species (ROS) and malondialdehyde (MDA) levels, while increasing glutathione (GSH) production in oxygen and glucose deprivation/reperfusion-induced PC12 cells. Transcriptome sequencing analysis of PC12 cells suggested that the protective effect of PTBP1KD-BMSCs against injury may involve ferroptosis. Furthermore, western blotting showed upregulation of glutathione synthetase (GSS), glutathione peroxidase 4 (GPX4), and solute carrier family 7 member 11 (SLC7A11) in PTBP1KD-BMSCs, known negative regulators of ferroptosis. Moreover, PTBP1KD-BMSCs inhibited p38MAPK and JNK activation. In addition, PTBP1KD-BMSCs transplantation into middle cerebral artery occlusion/reperfusion (MCAO/R) rats reduced cerebral infarction volume and improved neurological function. Immunofluorescence analysis confirmed the upregulation of GSS expression in neurons of the ischemic cortex, while immunohistochemistry indicated a downregulation of p-P38. These result suggest that PTBP1KD-BMSCs can alleviate neuronal IRI by reducing oxidative stress, inhibiting ferroptosis, and modulating the MAPK pathway, providing a theoretical basis for potential treatment strategies for cerebral IRI.

Ozone Administration Reduces Myocardial Ischemia Reperfusion Injury in Streptozotocin Induced Diabetes Mellitus Rat Model.

Objective: This study aimed to demonstrate whether ozone has cardioprotective effects on the myocardial ischemia-reperfusion injury (IRI) in rats with streptozotocin(STZ)-induced diabetes. Methods: A total of 38 male Wistar Albino rats were divided into five groups as follows: control group (group C,n=6), diabetic group (group D,n=6), diabetic ozone group (group DO,n=6), diabetic-ischemia/reperfusion (group DIR,n=6), diabetic-ischemia/reperfusion-ozone (group DIRO,n=6). Six rats died during this period and two died because of surgical complications. A myocardial ischemia-reperfusion model was created using a thoracotomy incision from 4th intercostal space. The LAD was ligated using an 8-0 prolene suture for 30min. Ozone was administered intraperitoneally(1mg/kg) 5min before reperfusion. The reperfusion time was 120 min. At the end of the reperfusion procedure, myocardial tissue histopathological examinations, and serum biochemical analyses were performed. Results: The percentage of TUNEL(+) cardiomyocytes/HPF was significantly higher in the DIR group than in the C, D, and DO groups. Conversely, TUNEL positivity was significantly lower in the DIRO group than in the DIR group. The IRI score was significantly higher in the DIR and DIRO groups than that in the C, D, and DO groups. In contrast, the IRI damage score in the DIRO group was significantly lower than that in the DIR group. Serum MDA levels were significantly higher in the DIR group than in the C, D, and DO groups. Similarly, MDA levels were significantly higher in the DIRO group than in the C and D groups. CAT activity was significantly higher in the DIR group than in the C and D groups. SOD activity was significantly higher in the DIR group than in the C and DO groups. Conclusion: Our study showed that ozone exerts cardioprotective effects in STZ-induced diabetic rats through its antioxidant role against oxidative stress. Both biochemical and histological analyses clearly revealed that ozone has beneficial effects against IRI in the diabetic rat myocardium.

Mitochondrial Function and Resistance to Oxidative Stress in the Kidney during Pregnancy.

We demonstrated that the serum of pregnant rats increases viability of kidney epithelial cells and promotes their proliferation. The intensity of oxidative stress in the kidneys was also reduced during pregnancy, but only in rats that were not exposed to acute ischemic kidney injury. This decrease in oxidative stress was not associated with changes in transmembrane mitochondrial potential, the size of mitochondria, time of opening of mitochondrial permeability transition pore (mPTP), mitochondrial respiration rate, antioxidant activity, or nitric oxide level.

Charge balanced aggregation: A universal approach to aqueous organic nanocrystals.

Organic nanocrystals, particularly those composed of conjugated molecules, hold immense potential for various applications. However, their practical utility is often hindered by the challenge of achieving stable aqueous dispersions, which are essential for biological compatibility and effective delivery. This study introduces a novel and versatile strategy for preparing stable aqueous organic nanocrystals using a modified reprecipitation method. We demonstrate the broad applicability of this approach by successfully preparing a diverse library of nanocrystals from 27 conjugated molecules. Our findings reveal a charge-balanced aggregation mechanism for nanocrystal formation, highlighting the crucial role of surface charge in controlling particle size and stability. Based on this mechanism, we establish a comprehensive molecular combination strategy that directly links molecular properties to colloidal behaviour, enabling the straightforward prediction and preparation of stable aqueous dispersions without the need for excipients. This strategy provides a practical workflow for tailoring the functionality of these nanocrystals for a wide range of applications. To illustrate their therapeutic potential, we demonstrate the enhanced efficacy of these nanocrystals in treating acute ulcerative colitis, myocardial ischemia/reperfusion injury, and cancer in mouse models. This work paves the way for developing next-generation nanomaterials with tailored functionalities for diverse biomedical applications.