The present study aimed to investigate the role of the C‑sis gene in the apoptosis of hepatocytes in vitro and in the liver function of a rat model of fulminant hepatic failure (FHF). Buffalo rat liver (BRL) cells were treated with hydrogen peroxide (H2O2) to induce apoptosis and then transfected with a C‑sis overexpression vector. A rat model of FHF was established, and C‑sis was overexpressed. The mRNA and protein expression of C‑sis were examined using reverse transcription‑polymerase chain reaction and western blot analyses, respectively. Cell viability was assessed by CCK8, and a TUNEL assay was used to examine cell apoptosis. Flow cytometry was used for cell cycle detection. Hematoxylin and eosin staining was used for histological examination. The levels of alanine transaminase (ALT) and aspartate transaminase (AST) were also examined in the rats. The results showed that C‑sis was successfully overexpressed in the cells and rat model. Compared with H2O2‑treated BRL cells, the overexpression of C‑sis significantly inhibited cell apoptosis, promoted cell viability, and decreased the expression of cleaved caspase-3. Similar results were observed in the FHF rats treated with the C‑sis overexpression plasmid, compared with those treated with empty plasmids. In addition, in the FHF rats overexpressing C‑sis, histological examination showed that liver injury was alleviated, the levels of ALT and AST were significantly decreased, and mortality rate was significantly decreased, compared with those observed in the rats treated with empty plasmids. In conclusion, the overexpression of C‑sis inhibited the H2O2‑induced apoptosis of BRL cells in vitro, and alleviated liver injury, improved liver function, and decreased mortality rates in rat models of FHF.

Myocardial injury is one of the main symptoms of sepsis. However, the mechanisms underlying sepsis‑induced myocardial dysfunction remain unclear. In the present study, the concentration of cardiac troponin T (CTnT) in serum was measured using an enzyme‑linked immunosorbent assay kit. The levels of interleukin (IL)‑1β and IL‑18 were assessed by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis and the level of malondialdehyde (MDA) was determined using a corresponding kit. Myocardial pathology was analyzed via hematoxylin and eosin staining. RT‑qPCR analysis and western blotting and/or immunohistochemistry were used to quantify the expression levels of thioredoxin‑interacting protein (TNXIP), NOD‑like receptor pyrin domain containing 3 (NLRP3), cleaved caspase‑1, caspase‑1, catalase and manganese‑superoxide dismutase (MnSOD). The viability of cells was determined using a cell counting kit‑8. Apoptosis and reactive oxygen species (ROS) were examined using flow cytometry. Models of sepsis‑induced myocardial injury were successfully established; evidence included increases in the levels of CTnT, IL‑1β, IL‑18 and MDA and myocardial tissue damage in vivo, and decreased cell viability and improvements in IL‑1β and IL‑18 in vitro. The levels of TXNIP, NLRP3 and cleaved caspase‑1 were upregulated in the sepsis models. Small interfering RNA targeting TNXNIP (siTXNIP) increased cell viability, reduced the apoptotic rate and attenuated the release of IL‑1β and IL‑18. The levels of TXNIP, NLRP3 and cleaved caspase‑1 and production of ROS were suppressed by siTXNIP, accompanied by increases in catalase and MnSOD. TXNIP/NLRP3 serves an important role in the development of sepsis‑induced myocardial damage.

Epithelial cell adhesion molecule (EpCAM) is highly expressed in mammalian intestines, and is essential for maintaining the homeostasis of the intestinal epithelium. EpCAM protein is localized at tight junctions and the basolateral membrane of the intestinal epithelium, where it interacts with many cell adhesion molecules. To explore the molecular functions of EpCAM in regulating adherens junctions in the intestinal epithelium, EpCAM knockout embryos and newborn pups were analyzed. Hematoxylin and eosin staining was used to assess the histology of the duodenum, jejunum, ileum and colon from wild-type and EpCAM‑/‑ mice at E18.5, P0 and P3. The expression and localization of adherens junction‑associated genes and genes that encode the proteins that participate in the assembly of adherens junctions were measured at the mRNA and protein levels using qPCR, western blot analysis and immunofluorescence staining. The results showed that although there was no significant damage to the intestines of EpCAM‑/‑ mice at E18.5 and P0, they were significantly damaged at P3 in mutant mice. The expression of adherens junction‑associated genes in EpCAM mutant mice was normal at the mRNA level from E18.5 to P3, but their protein levels were gradually reduced and mislocalized from E18.5 to P3. The expression of nectin 1, which can regulate the assembly and adhesion activity of E‑cadherin, was also gradually reduced at both the mRNA and protein levels in the intestinal epithelium of EpCAM mutant mice from E18.5 to P3. In summary, the loss of EpCAM may cause the reduction and mislocalization of proteins that compose adherens junctions partly via the downregulation of nectin 1 in the intestines.

The activation of mammalian ste20‑like kinase1 (Mst1) is a crucial event in cardiac disease development. The inhibition of Mst1 has been recently suggested as a potential therapeutic strategy for the treatment of diabetic cardiomyopathy. However, whether silencing Mst1 also protects against hypertensive (HP) myocardial injury, or the mechanisms through which this protection is conferred are not yet fully understood. The present study aimed to explore the role of Mst1 in HP myocardial injury using in vivo and in vitro hypertension (HP) models. Angiotensin II (Ang II) was used to establish HP mouse and cardiac microvascular endothelial cell (CMEC) models. CRISPR/adenovirus vector transfection was used to silence Mst1 in these models. Using echocardiography, hematoxylin and eosin staining, Masson's trichrome staining, the enzyme‑linked immunosorbent assay detection of inflammatory factors, the enzyme immunoassay detection of oxidative stress markers, terminal deoxynucleotidyl transferase dUTP nick‑end labeling staining, scanning electron microscopy, transmission electron microscopy, as well as immunofluorescence and western blot analysis of the autophagy markers, p62, microtubule‑associated proteins 1A/1B light chain 3B and Beclin‑1, it was found that Ang II induced HP myocardial injury with impaired cardiac function, increased the expression of inflammatory factors, and elevated oxidative stress in mice. In addition, it was found that Ang II reduced autophagy, enhanced apoptosis, and disrupted endothelial integrity and mitochondrial membrane potential in cultured CMECs. The silencing of Mst1 in both in vivo and in vitro HP models attenuated the HP myocardial injury. On the whole, these findings suggest that Mst1 is a key contributor to HP myocardial injury through the regulation of cardiomyocyte autophagy.

A number of macrophage phenotypes have been previously identified as crucial regulators in the progression of hepatic fibrosis (HF). Cytokines from macrophages or Kupffer cells (KCs) have also been identified to be important regulators in HF. Blocking Kv1.3 in models of HF, regulating macrophage polarization and cytokine secretion have not yet been assessed as potential treatments options for this condition. In the current study, a model of carbon tetrachloride (CCl4)‑induced HF was established and examined the effects of margatoxin (MgTX; an inhibitor of Kv1.3) on HF. Hematoxylin and eosin, Masson's trichrome and immunohistochemistry staining were performed to determine whether MgTX can alleviate liver fibrosis. To elucidate the mechanisms through which MgTX attenuates liver injury, reverse transcription‑quantitative PCR and western blot analysis were used to detect polarized macrophage markers in RAW264.7 cells and cytokines were examined using ELISA. Furthermore, macrophage polarization signal transducer and activator of transcription (STAT) signaling, which is associated with macrophage polarization, was identified in RAW264.7 cells. The results revealed that MgTX protected the mice from CCl4‑induced liver fibrosis. Furthermore, MgTX decreased the expression of M1 phenotype biomarkers, and increased the expression of M2 phenotype biomarkers in CCl4‑induced HF. Additionally, the production of pro‑inflammatory cytokines was decreased and interleukin‑10 production was increased in the serum of mice with HF injected with MgTX. Furthermore, MgTX was found to regulate the expression of M1 markers by suppressing p‑STAT1 activity and increasing the expression of M2 markers by promoting p‑STAT6 activity. On the whole, the findings of this study demonstrate that MgTX is able to alleviate CCl4‑induced HF in mice, possibly via macrophage polarization, cytokine secretion and STAT signaling.

In order to explore the potential effects of interleukin (IL)-35 on IL-10, transforming growth factor-β (TGF-β), interferon-γ (INF)-γ, IL-12 and IL-17, a pcDNA3.1‑IL-35 plasmid was injected into the vitreous cavity of BALB/c mice. Enzyme-linked immunosorbent assay, western blot analysis and quantitative PCR analysis were performed to confirm the successful expression of IL-35. Slit-lamp biomicroscopy, hematoxylin and eosin staining and immunofluorescence were employed to detect the status of eyes, and western blot analysis was performed to examine the expression of corneal graft rejection-related cytokines. There were no abnormalities in the eyes pre-mydriasis or post-mydriasis and no injuries to the cornea or retina following the injection of IL-35-expressing plasmid. An immunofluorescence assay detected the positive expression of IL-35 in corneal epithelial cells from IL-35‑injected mice and negative staining in the control group. Further study revealed that IL-35 enhanced the expression of IL-10 and TGF-β which reached their highest levels at 1 and 2 weeks after injection, respectively (p<0.01). Moreover, the expression of INF-γ and IL-12 was decreased significantly at 2 weeks after the injection of IL-35-expressing plasmid (p<0.05), and the expression of IL-17 was suppressed notably at 4 weeks after the injection (p<0.05). The intravitreal injection of IL-35-expressing plasmid in mice downregulates the expression of pro-inflammatory cytokines and upregulates the expression of anti-inflammatory cytokines. Thus, IL-35 may further be assessed as a potential target for the treatment of corneal graft rejection.

Hesperidin has been reported to attenuate myocardial ischemia/reperfusion (I/R) injury; however, its effect on autophagy during myocardial I/R and the underlying mechanism remains unknown. The present study aimed to investigate whether hesperidin inhibited I/R‑induced excessive myocardial autophagy through activating the phosphatidylinositol 3‑kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway. Male adult rats were pretreated with hesperidin for a total of 3 days prior to ischemia in the absence or presence of LY294002, a PI3K inhibitor, and then subjected to ischemia for 30 min followed by reperfusion for 4 h. Myocardial infarct size was measured by Evans blue/triphenyltetrazolium chloride staining. Hematoxylin and eosin staining was used for observing the histological changes in the heart, and the serum levels of creatine kinase‑MB (CK‑MB) and cardiac troponin I (cTnI) were measured by enzyme‑linked immunosorbent assay. Additionally, the protein levels of light chain (LC) 3Ⅱ, Beclin1, phosphorylated (p)‑mTOR, p‑Akt and p‑PI3K were determined by western blot analysis. Hesperidin pretreatment significantly decreased the myocardial infarct size, myocardial damage and serum levels of CK‑MB and cTnI. Furthermore, the expression levels of LC3Ⅱ and Beclin1 were significantly downregulated and the expression levels of p‑mTOR, p‑Akt and p‑PI3K were markedly upregulated by hesperidin. However, the aforementioned effects as a result of hesperidin were significantly reversed by the presence of LY294002. These results demonstrated that hesperidin reduced myocardial I/R injury by suppressing excessive autophagy. Activation of the PI3K/Akt/mTOR pathway contributed to the inhibitory effect of hesperidin on excessive autophagy.

MicroRNAs (miRNAs) play an important role in the development of vascular remodeling in essential hypertension (EH) by mediating the effects of human cytomegalovirus (HCMV) on the vascular system. Therefore, the aim of the present study was to investigate the effects of murine cytomegalovirus (MCMV) infection on blood pressure and vascular function in mice, in order to elucidate the role of miR‑1929‑3p in this process. For model development, 7‑month‑old C57BL/6J mice were infected with the Smith strain of MCMV, and MCMV DNA, IgG and IgM were detected. Subsequently, blood pressure was measured via the carotid artery, and the morphological changes of the aorta were evaluated by hematoxylin and eosin and Masson's trichrome staining. miR‑1929‑3p transfection was performed using an adeno‑associated virus packaged vector and the changes in vascular structure were then observed. The levels of nitric oxide (NO) and endothelial NO synthase were also assessed with colorimetry. Vascular remodeling and expression of NLRP3 inflammasome pathway‑related proteins were detected by immunohistochemistry and western blotting. Endothelin‑1 (ET‑1), interleukin (IL)‑1β and IL‑18 were assayed by ELISA. The results revealed that MCMV infection increased the blood pressure, promoted vascular remodeling, caused endothelial cell injury, and downregulated miR‑1929‑3p. However, these effects were alleviated by miR‑1929‑3p overexpression, which downregulated endothelin A receptor (Ednra) and NLRP3 inflammasome, as well as endothelial injury‑ and vascular remodeling‑related genes. Taken together, the findings of the present study indicated that overexpression of miR‑1929‑3p may improve MCMV‑induced vascular remodeling, possibly via the deactivation of the NLRP3 inflammasome by ET‑1/Ednra.

Non‑alcoholic fatty liver disease (NAFLD) is the consequence of insulin resistance, fatty acid accumulation, oxidative stress and lipotoxicity. The present study aimed to elucidate the effect of Quaking 5 (QKI 5) as mediated by Sirtuin 1 (SIRT1) on triglyceride (TG) synthesis in the liver of an NAFLD mouse model. A high‑fat diet‑induced NAFLD model was established in mice, and mouse hepatocytes were isolated to characterize the effects of QKI 5 mediated by SIRT1 on TG synthesis in the liver. Body weight and liver wet weight were recorded. In addition, serum levels of total cholesterol, TG, alanine aminotransferase and aspartate aminotransferase were assessed using an automatic biochemistry analyzer. Hematoxylin and eosin staining was performed to observe the histological morphological alterations of the liver tissues. The concentration of SIRT1 in the serum was also detected. The NAFLD activity score (NAS) was used to evaluate disease severity. The synthesis of TGs in cells or tissues was determined, and the protein levels of SIRT1, QKI 5, peroxisome proliferator‑activated receptor (PPAR)α and Forkhead box protein O1 (FoxO1) were examined. The expression levels of SIRT1 or QKI 5, and the acetylation level of QKI 5 were decreased in the mouse model of NAFLD. QKI 5 was deacetylated by SIRT1, which contributed in suppressing the progression of NAFLD in the mice, and inhibiting TG synthesis in vivo and in vitro via the PPARα/FoxO1 signaling pathway. Taken together, the results of the present study demonstrated that SIRT1 deacetylated QKI 5, an RNA‑binding protein significantly affecting the synthesis of TG in the liver of the NAFLD mouse model. Furthermore, it activated transcription factor FOXO1 through post‑transcriptional regulation of the expression of PPARα and further inhibited the synthesis of TGs, thereby restraining the progression of NAFLD.

Type 2 diabetes mellitus (T2DM) is associated with chronic low‑grade inflammation. Carvacrol has been confirmed to possess anti‑inflammatory properties, but its effect on diabetic vasculature remains unknown. The aim of the present study was to investigate the possible protective effects of carvacrol against vascular endothelial inflammation. The mice were divided into four groups (n=15 per group) as follows: Non‑diabetic control mice, db/db mice, db/db mice + carvacrol (low) and db/db mice + carvacrol (high) groups. The effects of carvacrol on the pathomorphism of the thoracoabdominal aorta in db/db mice were evaluated using hematoxylin and eosin and Masson's trichrome staining. The serum levels of insulin signaling molecules, such as phosphorylated insulin receptor, phosphorylated insulin receptor substrate‑1, insulin, triglyceride (TG) and inflammatory cytokines [tumor necrosis factor‑α, interleukin (IL)‑1β, IL‑6 and IL‑8] were measured by ELISA. Furthermore, the protein levels of the toll‑like receptor (TLR)4/nuclear factor (NF)‑κB inflammatory signaling pathway molecules were investigated in the thoracoabdominal aorta of db/db mice and in high glucose‑induced endothelial cells. Vascular endothelial cell apoptosis and viability were assessed by using flow cytometry and Cell Counting Kit‑8 assays, respectively. The results demonstrated that carvacrol alleviated vascular endothelial cell injury. Carvacrol reduced the expression levels of insulin signaling molecules, insulin, TG and inflammatory cytokines in the serum of db/db mice. Moreover, carvacrol reduced the activation of the TLR4/NF‑κB signaling pathway in vivo and in vitro. In vitro, carvacrol inhibited high glucose‑induced endothelial cell function by promoting vascular endothelial cell apoptosis and suppressing cell viability. These findings demonstrated that carvacrol could alleviate endothelial dysfunction and vascular inflammation in T2DM.

Myocardial ischemia/reperfusion (I/R) induces cardiac cell injury; however, the mechanism underlying cardiac damage remains unclear. A previous study demonstrated that triptolide (TP) exerts protective effects against I/R in cerebral cells. The present study aimed to evaluate the protective effects of TP on cardiac cells, and investigated the potential mechanisms involved in I/R‑induced damage. Rats and cardiac H9C2 cells undergoing I/R were pretreated with TP, and cell damage was assessed in vivo and in vitro. Hematoxylin and eosin and terminal deoxynucleotidyl‑transferase‑mediated dUTP nick end labeling staining were employed to evaluate I/R injury in rat cardiac tissue. Inflammatory factors, including tumor necrosis factor‑α, interleukin (IL)‑1β and IL‑6, were detected by ELISA. Biochemical analyses were performed to evaluate the bioactivity of superoxide dismutase, malondialdehyde and catalase. In addition, viability of H9C2 cells was measured using the Cell Counting kit 8 assay. Flow cytometry was used to evaluate cell apoptosis and reactive oxygen species (ROS) generation. Furthermore, the expression levels of proteins associated with apoptosis, peroxide and inflammation were measured using western blot analysis. H9C2 cells were also treated with N‑acetylcysteine and pyrrolidine dithiocarbamate, and cell injury was assessed after peroxidation or I/R. The results demonstrated that TP exerted a significant protective effect on cardiac cells in vivo and in vitro. TP reduced the inflammatory response, as determined by nuclear factor‑κB inhibition. In addition, TP decreased ROS‑mediated lipid peroxidation, and reduced ROS generation. TP also inhibited cell apoptosis by activating the extracellular signal‑regulated kinase 1/2 pathway. In conclusion, TP may protect cardiac cells from I/R injury; the potential protective mechanisms of TP against I/R include anti‑inflammatory action, antioxidation and apoptotic resistance.

The inhibition of mesangial cell (MC) proliferation has become an important therapy in preventing glomerular proliferation diseases. Trifluoperazine (TFP) has been reported to inhibit the proliferation of several types of cancer cell, however, the effects of TFP in renal proliferation diseases remain to be fully elucidated. The present study examined the effects of TFP on the proliferation of MCs and quantified cell apoptosis progression in vivo and in vitro. The effects of various TFP concentrations and treatment durations on cell proliferation and cell apoptosis in vitro were analyzed using flow cytometry in conjunction with a Cell Counting kit‑8 assay. Cell proliferation in vivo was determined using hematoxylin and eosin staining and immunohistochemistry of Ki67. The expression of the two cell apoptosis‑related proteins, B‑cell lymphoma-2 (Bcl‑2) and Bcl‑2‑associated X protein (Bax), were estimated using western blot analysis and immunohistochemistry in vivo and in vitro. TFP‑induced phosphatidylinositol 3‑kinase (PI3K)/protein kinase B (AKT) signaling pathways were also estimated using western blot analysis. These results suggested that TFP inhibited MC proliferation in a dose‑ and time‑dependent manner. It was found that TFP inhibited the abnormal proliferation of MCs, which was stimulated by 20% fetal bovine serum in vitro and in lupus MRL/lpr mice. TFP promoted cell apoptosis, downregulated the expression of Bcl‑2 and upregulated the expression of Bax in a dose‑dependent manner at mRNA and protein levels. In addition, TFP inhibited phosphorylated AKT, potentially leading to the suppressed activation of PI3K/AKT signaling pathways. TFP treatment significantly decreased the levels of blood urea nitrogen and serum creatinine, but had no significant effects on the body weight and liver function of the lupus mice. These results validated and reinforced the potential of TFP in the treatment of mesangial proliferative diseases.

The aim of the present study was to elucidate the anti-inflammatory effects of the two novel anti-inflammatory substances, 3-[(dodecylthiocarbonyl)‑methyl]-glutarimide (DTCM-G) and dehydroxymethylepoxyquinomicin (DHMEQ), on DSS-induced colitis in rats. For this purpose, rats with dextran sulfate sodium (DSS)-induced colitis were randomly divided into 3 groups with 10 animals in each group as follows: i) the control group, which received 0.5 ml of 0.5% carboxymethyl cellulose (CMC; vehicle), ii) rats that received DTCM-G (20 mg/kg body weight in 0.5% CMC; the DTCM-G group), and iii) rats that received DHMEQ (15 mg/kg body weight in 0.5% CMC; the DHMEQ group). The animals were sacrificed after the 5-day treatment period, and tissue samples were taken from their colons and sectioned for histological evaluation. The tissue sections were stained with hematoxylin and eosin, and immunostained for leukocytes, lymphocytes, macrophages/monocytes and mast cells. The disease activity index (DAI), histological grading of colitis, and densities of several types of submucosal immune cells were compared between the controls, and the DTCM-G and DHMEQ groups. The DAI values were significantly lower in both the DTCM-G and DHMEQ groups than in the control group. The total scores for the histological grading of colitis were also significantly lower in the DTCM-G and DHMEQ groups than in the control group. The submucosal densities of leucocytes, lymphocytes, macrophages/monocytes and mast cells were significantly lower in the DTCM-G and DHMEQ groups than in the control group. Our findings indicate that the anti-inflammatory and anticancer effects of DTCM-G and DHMEQ, and the absence of any associated toxicity render them excellent therapeutic candidates for clinical use in the treatment of colitis.

Hepatic stellate cells (HSCs) are the main extracellular matrix (ECM)‑producing cells in liver fibrosis. Activated HSCs stimulate the proliferation and migration of hepatocellular carcinoma (HCC) cells. Cysteine‑rich 61 (CCN1/Cyr61) is an ECM protein. Our previous studies demonstrated that the expression of CCN1 was significantly higher in benign hepatic cirrhosis tissue and cancer‑adjacent hepatic cirrhosis tissues. CCN1 is a target gene of β‑catenin in HCC and promotes the proliferation of HCC cells. The present study aimed to examine whether CCN1 can activate HSCs and affect the function of activated HSCs in promoting the progression of HCC. CCN1 expression was determined during the progression of liver fibrosis in a mouse model. LX‑2 cells, which were infected with adenoviruses AdCCN1 or AdRFP, and HepG2 cells were co‑cultured or subcutaneously co‑implanted into in nude mice. MTT assay, Crystal Violet staining, Boyden chamber, matrigel invasion and monolayer scratch assays were used to analyze the proliferation, migration and invasion capability of HepG2 cells. Xenograft sizes were measured and histological analyses were performed by hematoxylin and eosin, immunohistochemical, immunefluorescence and Sirius Red staining. It was demonstrated that the expression of CCN1 was continually increased in liver fibrosis and the that expression may be correlated with the progression of liver fibrosis. CCN1 affected the function of LX‑2 and enhanced the effect of LX‑2 on promoting the viability, migration and invasion of HepG2 cells in vitro. CCN1 enhanced the effect of LX‑2 on promoting the growth of HepG2 xenografts in vivo. CCN1 also affected the function of activated HSCs and regulated the formation of the xenograft microenvironment, including fibrogenesis and angiogenesis, which are beneficial for the progression of HCC. These findings demonstrated that CCN1 may be involved in the progression of the hepatic cirrhosis‑HCC axis through regulating HSCs.

Ischemic postconditioning (IPoC) has been demonstrated to prevent myocardial ischemia‑reperfusion injury (MIRI), but its cardioprotective effect is abrogated by hypercholesterolemia. The aim of the present study was to determine whether lycopene (LP), a type of carotenoid, can restore the cardioprotective effect of IPoC in hypercholesterolemic rats. Male Wistar rats were fed a cholesterol‑enriched diet for 12 weeks to establish a hypercholesterolemic model. The rat hearts were isolated and subjected to 30 min ischemia and 60 min reperfusion using a Langendorff apparatus. LP was administered to the rats intraperitoneally for 5 consecutive days prior to ischemia and reperfusion. Myocardial pathological changes, infarct size and cell apoptosis were measured by hematoxylin and eosin, triphenyltetrazolium chloride and TUNEL staining, respectively. The changes in endoplasmic reticulum (ER) stress markers, the reperfusion injury salvage kinase (RISK) pathway and mitochondrial apoptosis‑related proteins were detected by western blotting. Overall, the results demonstrated that low‑dose LP in combination with IPoC ameliorated myocardial histopathological changes, reduced the infarct size and release of cardiac enzymes, and decreased cardiomyocyte apoptosis in hypercholesterolemic rats, but no beneficial effects were achieved by the same dose of LP or IPoC treatment were used alone. Furthermore, the combination of LP and IPoC inhibited the expression of glucose‑regulated protein 78 and C/EBP homologous protein, increased the phosphorylation levels of AKT, ERK1/2 and glycogen synthase kinase‑3β, repressed mitochondrial permeability transition pore opening, and reduced the expression of cytochrome c, cleaved caspase‑9 and cleaved caspase‑3. Collectively, these findings demonstrated that LP can restore the cardioprotective effects of IPoC on MIRI in hypercholesterolemic rats, and this restoration by LP was mediated by inhibition of ER stress and reactivation of the RISK pathway in hypercholesterolemic rat myocardium.

Raf kinase inhibitor protein (RKIP) is an inflammation‑inhibiting mediator that is involved in several diseases; however, the potential mechanism of action of RKIP on the inflammatory response induced by influenza A virus (IAV) remains unclear. The present study aimed to investigate whether RKIP regulated the inflammatory response via the ERK/MAPK pathway. The present study detected the expression levels of RKIP and alterations in the inflammatory response in human normal bronchial epithelial BEAS‑2B cells, diseased human bronchial epithelial cells and primary human bronchial epithelial cells infected with IAV. Cells were treated with locostatin to inhibit the expression of RKIP. RKIP was overexpressed by lentivirus transduction and the small molecule inhibitor SCH772984 was applied to specifically inhibit activation of the ERK/MAPK pathway. In addition, C57BL/6 mice were infected with IAV to further confirm the role of RKIP in regulation of the inflammatory response via ERK/MAPK in vivo. Western blotting, reverse transcription‑quantitative PCR, ELISA, 5‑ethynyl‑-2'‑deoxyuridine assay, immunofluorescence staining, Cell Counting Kit‑8, cell cycle assay, hematoxylin and eosin staining, and immunohistochemistry were used to detect all of the changes. Notably, RKIP attenuated the inflammatory response that was triggered by IAV infection in airway epithelial cells, which was characterized by augmented inflammatory cytokines and cell cycle arrest. Furthermore, the ERK/MAPK pathway was revealed to be activated by IAV infection and downregulation of RKIP aggravated the airway inflammatory response. By contrast, overexpression of RKIP effectively ameliorated the airway inflammatory response induced by IAV. These findings demonstrated that RKIP may serve a protective role in airway epithelial cells by combating inflammation via the ERK/MAPK pathway. Collectively, the present findings suggested that RKIP may negatively regulate airway inflammation and thus may constitute a promising therapeutic strategy for airway inflammatory‑related diseases that are induced by IAV.

The mechanism underlying the anti‑inflammatory or antifibrotic activity of erythropoietin (EPO) in myocardial fibrosis (MF) remains elusive. In the current study, abdominal aortic constriction (AAC) was performed on rats and EPO and/or Toll‑like receptor (TLR)4 were overexpressed in rat hearts through intramyocardial administration of lentivirus expressing the EPO and TLR4 genes. Hematoxylin and eosin staining and Masson's trichrome staining were performed on tissue sections from rat hearts for histopathological examination. ELISA was used to determine the levels of inflammatory mediators in serum. Gene expression levels were determined by quantitative polymerase chain reaction analysis and protein expression levels were determined by western blot analysis and immunofluorescence staining. The results indicated that EPO overexpression improved MF in rat hearts, by inhibiting the release of transforming growth factor (TGF)‑β1, tumor necrosis factor (TNF)‑α, interleukin (IL)‑6, IL‑1β, IL‑17A, matrix metalloproteinase (MMP)‑9 and MMP‑2. Moreover, EPO overexpression suppressed the expression of TLR4, while promoting phosphoinositide 3‑kinase (PI3K) and phosphorylated AKT serine/threonine kinase 1 (Akt) expression levels. However, the beneficial effects of EPO were attenuated by overexpression of TLR4. In addition, inhibition of PI3K/Akt signaling activity by treatment with LY294002 markedly reversed the protective effect of EPO on the AAC‑induced MF. Taken together, the present study demonstrated that EPO may have a critical role against MF by activating PI3K/Akt signaling and by downregulating TLR4 expression, thereby inhibiting the release of TGF‑β1, TNF‑α, IL‑6, IL‑1β, IL‑17A, MMP‑9 and MMP‑2. These findings suggest that the PI3K/Akt/TLR4 signaling pathway is associated with the anti‑inflammatory effects of EPO and may play a role in attenuating AAC‑induced MF.

Preeclampsia (PE) is a disorder that is characterized by pregnancy‑induced hypertension. It has been reported that Annexin A1 (ANXA1) is highly expressed in the plasma of women diagnosed with PE. Therefore, the present study aimed to examine the effect of ANXA1 on PE rats. The PE animal model was constructed in rats using Nω‑nitro‑L‑arginine methyl ester (L‑NAME), and the blood pressure and urine protein levels of rats were detected. The pathological features of placental tissue, and the levels of inflammatory factors and ANXA1 were respectively measured by hematoxylin‑eosin staining, enzyme‑linked immunosorbent assay and immunohistochemical assay. The activity of trophoblasts obtained from PE placental tissue was measured using immunofluorescence staining, while cell apoptosis was assessed using flow cytometry. The levels of associated factors were determined by reverse transcription‑quantitative polymerase chain reaction and western blot analysis. The results identified that systolic blood pressure, diastolic blood pressure, mean arterial pressure and urine protein levels were enhanced, and that the contents of ANXA1, tumor necrosis factor alpha (TNF‑α), interleukin (IL)‑1β, IL‑6 and IL‑8 were increased in the L‑NAME group. Transfection with small interfering RNA (siRNA)‑ANXA1 markedly decreased the apoptosis and inflammatory response of trophoblasts. In addition, siRNA‑ANXA1 upregulated the levels of B‑cell lymphoma‑2 (Bcl‑2) and pro‑caspase‑3, and downregulated the levels of Bcl‑2‑associated X protein, cleaved‑caspase‑3, TNF‑α, IL‑1β, IL‑6 and IL‑8. Furthermore, siRNA‑ANXA1 repressed the phosphorylation of Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3); however, siRNA‑ANXA1 did not alter the levels of JAK2 and STAT3. Therefore, silencing of ANXA1 suppressed the apoptosis and inflammatory response of PE rat trophoblasts, and downregulated JAK2/STAK3 pathway.

The present study aimed to investigate the protective effects of rhein on cerebral ischemic/reperfusion (I/R) injury in rats. The present study focused on the effect of rhein on oxidative stress and apoptotic factors, which are considered to serve an important role in the onset of I/R injury. Sprague‑Dawley rats were subjected to middle cerebral artery occlusion. Neurological functional scores (NFSs) were evaluated according to the Zea Longa's score criteria and the area of brain infarct was determined by triphenyltetrazolium chloride staining. The morphology of the nerve cells in the cortex was observed following hematoxylin and eosin staining. In addition, levels of oxidative stress were assessed by measuring the levels of superoxide dismutase (SOD), glutathione‑peroxidase (GSH‑Px), catalase (CAT) and malondialdehyde (MDA). Levels of B‑cell lymphoma-2 (Bcl‑2), apoptosis regulator Bax (BAX), caspase-9, caspase‑3 and cleaved caspase‑3 expression were analyzed using western blot analysis. Levels of caspase‑9 and caspase‑3 mRNA expression were obtained using reverse transcription‑quantitative polymerase chain reaction. The results revealed that treatment with 50 or 100 mg/kg rhein significantly improved the NFS and markedly attenuated the area of infarction. Rhein also significantly reduced the content of MDA and significantly increased SOD, GSH‑Px and CAT activity. Western blot analysis indicated that rhein significantly decreased the expression of BAX and enhanced the expression of Bcl‑2. Compared with the I/R group, levels of caspase‑9, caspase‑3 and cleaved caspase‑3 protein expression were significantly decreased in the rhein treatment groups. Additionally, rhein treatment significantly reduced levels of caspase‑9 and caspase‑3 mRNA expression. These results suggest that rhein exhibits protective effects during cerebral I/R injury and its underlying mechanism of action may involve the inhibition of oxidative stress and apoptosis.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are severe clinical conditions with a high mortality rate. Nucleotide‑binding oligomerization domain (NOD)‑like receptor containing pyrin domain 3 (NLRP3) and nuclear factor E2‑related factor 2 (Nrf2) have been reported to be associated with ALI. However, the dynamic changes in the levels of these factors in lipopolysaccharide (LPS)‑induced lung injury remain unclear. Thus, the present study aimed to determine the LPS‑induced activation of immunological cascades, as well as the NLRP3/Nrf2 signaling pathway at different stages of lung injury. For this purpose, mice were divided into six groups as follows: The control, LPS‑4 h, LPS‑24 h, LPS‑48 h, LPS‑96 h and LPS‑144 h groups. LPS (4 mg/kg) was administered intratracheally to induce lung injury. Flow cytometry was used to determine the changes in macrophages, neutrophils and T‑cell subsets in lung tissue, hematoxylin and eosin staining were used to measure the histopathological changes in lung tissues, ELISA was performed to evaluate the levels of cytokines, western blot analysis was used to measure the levels of inflammatory proteins, and reverse transcription‑quantitative PCR used to determine the mRNA level of a target gene. Following LPS administration, evident histopathological damage with neutrophil infiltration was observed which peaked at 48 h. The levels of interleukin‑1β, keratinocyte‑derived chemokine, macrophage inflammatory protein 2 and tumor necrosis factor a were markedly increased in bronchoalveolar lavage fluid and serum from the mice, and these levels peaked at 4 h. Moreover, LPS promoted Toll like receptor‑4 expression and reactive oxygen species production, thus activating NLRP3/Nrf2 signaling and pyroptosis. Collectively, the present study demonstrates that LPS triggers multiple inflammatory molecules and immune cells during ALI, which may be closely involved in the irregular redox status, NLRP3/Nrf2 pathway and pyroptosis.