Xijiao Dihuang Decoction Protects Against Murine Sepsis-Induced Cardiac Inflammation and Apoptosis via Suppressing TLR4/NF-κB and Activating PI3K/AKT Pathway.

Background: Xijiao Dihuang decoction (XJDHT), a traditional Chinese medicine, is widely used to treat patients with sepsis. However, the mechanisms underlying the effects of XJDHT on cardiac dysfunction have yet to be fully elucidated. The present study evaluated the potential utility of XJDHT in protecting against sepsis-induced cardiac dysfunction and myocardial injury. Methods: The mice were randomly divided into 3 groups and administered Lipopolysaccharide (LPS,10 mg/kg) or equivalent saline solution (control) and treated with XJDHT (10 g/kg/day) or saline by gavage for 72 hours. XJDHT was dissolved in 0.9% sodium chloride and administered at 200 µL per mouse. Transthoracic echocardiography, RNA-seq, TUNEL assays and hematoxylin and eosin (H&E) staining of cardiac tissues were performed. Results: Treatment with XJDHT significantly enhanced myocardial function and attenuated pathological change, infiltration of inflammatory cells, levels of TNF-α, IL-1ß and expression of TLR4 and NF-κB in mice with sepsis. RNA sequencing and Kyoto Encyclopedia of Genes and Genomes pathway analyses identified 531 differentially expressed genes and multiple enriched signaling pathways including the PI3K/AKT pathway. Further, XJDHT attenuated cardiac apoptosis and decreased Bax protein expression while increasing protein levels of Bcl-2, PI3K, and p-AKT in cardiac tissues of mice with sepsis. Conclusion: In summary, XJDHT improves cardiac function in a murine model of sepsis by attenuating cardiac inflammation and apoptosis via suppressing the TLR4/NF-κB pathway and activating the PI3K/AKT pathway.

Qingda granule ameliorates vascular remodeling and phenotypic transformation of adventitial fibroblasts via suppressing the TGF-ß1/Smad2/3 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingda granule (QDG) exhibits significant therapeutic effects on high blood pressure, vascular dysfunction, and elevated proliferation of vascular smooth muscle cells by inhibiting multiple pathways. However, the effects and underlying mechanisms of QDG treatment on hypertensive vascular remodeling are unclear. AIM OF THE STUDY: The aim of this study was to determine the role of QDG treatment in hypertensive vascular remodeling in vivo and in vitro. MATERIALS AND METHODS: An ACQUITY UPLC I-Class system coupled with a Xevo XS quadrupole time of flight mass spectrometer was used to characterize the chemical components of QDG. Twenty-five spontaneously hypertensive rats (SHR) were randomly divided into five groups, including SHR (equal volume of double distilled water, ddH2O), SHR + QDG-L (0.45 g/kg/day), SHR + QDG-M (0.9 g/kg/day), SHR + QDG-H (1.8 g/kg/day), and SHR + Valsartan (7.2 mg/kg/day) groups. QDG, Valsartan, and ddH2O were administered intragastrically once a day for 10 weeks. For the control group, ddH2O was intragastrically administered to five Wistar Kyoto rats (WKY group). Vascular function, pathological changes, and collagen deposition in the abdominal aorta were evaluated using animal ultrasound, hematoxylin and eosin and Masson staining, and immunohistochemistry. Isobaric tags for relative and absolute quantification (iTRAQ) was performed to identify differentially expressed proteins (DEPs) in the abdominal aorta, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed. Cell Counting Kit-8 assays, phalloidin staining, transwell assays, and western-blotting were performed to explore the underlying mechanisms in primary isolated adventitial fibroblasts (AFs) stimulated with transforming growth factor-ß 1 (TGF-ß1) with or without QDG treatment. RESULTS: Twelve compounds were identified from the total ion chromatogram fingerprint of QDG. In the SHR group, QDG treatment significantly attenuated the increased pulse wave velocity, aortic wall thickening, and abdominal aorta pathological changes and decreased Collagen I, Collagen III, and Fibronectin expression. The iTRAQ analysis identified 306 DEPs between SHR and WKY and 147 DEPs between QDG and SHR. GO and KEGG pathway analyses of the DEPs identified multiple pathways and functional processes involving vascular remodeling, including the TGF-ß receptor signaling pathway. QDG treatment significantly attenuated the increased cell migration, actin cytoskeleton remodeling, and Collagen I, Collagen III, and Fibronectin expression in AFs stimulated with TGF-ß1. QDG treatment significantly decreased TGF-ß1 protein expression in abdominal aortic tissues in the SHR group and p-Smad2 and p-Smad3 protein expression in TGF-ß1-stimulated AFs. CONCLUSIONS: QDG treatment attenuated hypertension-induced vascular remodeling of the abdominal aorta and phenotypic transformation of adventitial fibroblasts, at least partly by suppressing TGF-ß1/Smad2/3 signaling.

Performance assessment of medical service for organ transplant department based on diagnosis-related groups: A programme incorporating ischemia-free liver transplantation in China.

Background: In July 2017, the first affiliated hospital of Sun Yat-sen university carried out the world's first case of ischemia-free liver transplantation (IFLT). This study aimed to evaluate the performance of medical services pre- and post-IFLT implementation in the organ transplant department of this hospital based on diagnosis-related groups, so as to provide a data basis for the clinical practice of the organ transplant specialty. Methods: The first pages of medical records of inpatients in the organ transplant department from 2016 to 2019 were collected. The China version Diagnosis-related groups (DRGs) were used as a risk adjustment tool to compare the income structure, service availability, service efficiency and service safety of the organ transplant department between the pre- and post-IFLT implementation periods. Results: Income structure of the organ transplant department was more optimized in the post-IFLT period compared with that in the pre-IFLT period. Medical service performance parameters of the organ transplant department in the post-IFLT period were better than those in the pre-IFLT period. Specifically, case mix index values were 2.65 and 2.89 in the pre- and post-IFLT periods, respectively (p = 0.173). Proportions of organ transplantation cases were 14.16 and 18.27%, respectively (p < 0.001). Compared with that in the pre-IFLT period, the average postoperative hospital stay of liver transplants decreased by 11.40% (30.17 vs. 26.73 days, p = 0.006), and the average postoperative hospital stay of renal transplants decreased by 7.61% (25.23 vs.23.31 days, p = 0.092). Cost efficiency index decreased significantly compared with that in the pre-IFLT period (p < 0.001), while time efficiency index fluctuated around 0.83 in the pre- and post-IFLT periods (p = 0.725). Moreover, the average postoperative hospital stay of IFLT cases was significantly shorter than that of conventional liver transplant cases (p = 0.001). Conclusion: The application of IFLT technology could contribute to improving the medical service performance of the organ transplant department. Meanwhile, the DRGs tool may help transplant departments to coordinate the future delivery planning of medical service.

Ribosome assembly factor PNO1 is associated with progression and promotes tumorigenesis in triple­negative breast cancer.

The aim of the present study was to investigate the expression of ribosome assembly factor partner of NOB1 homolog (PNO1) and its association with the progression of breast cancer (BC) in patients, as well as its biological function and underlying mechanism of action in BC cells. Bioinformatics and immunohistochemical analyses revealed that PNO1 expression was significantly increased in BC tissues and its high mRNA expression was associated with shorter overall survival (OS) and relapse­free survival (RFS) of patients with BC, as well as multiple clinical characteristics (including advanced stage of NPI and SBR, etc.) of patients with BC. Biological functional studies revealed that transduction of lentivirus encoding sh­PNO1 significantly downregulated PNO1 expression, reduced cell confluency and the number of BC cells in vitro and inhibited tumor growth in vivo. Moreover, PNO1 knockdown decreased the cell viability and arrested cell cycle progression at the G2/M phase, as well as downregulated cyclin B1 (CCNB1) and cyclin­dependent kinase 1 (CDK1) protein expression in BC cells. Correlation analysis demonstrated that PNO1 expression was positively correlated with both CDK1 and CCNB1 expression in BC samples. Collectively, PNO1 was upregulated in BC and associated with BC patient survival, and PNO1 knockdown suppressed tumor growth in vitro and in vivo. In addition, positive regulation of CCNB1 and CDK1 may be one of the underlying mechanisms.

Uncaria Rhynchophylla attenuates angiotensin â ¡-induced myocardial fibrosis via suppression of the RhoA/ROCK1 pathway.

Uncaria rhynchophylla (UR), a traditional Chinese medicine, has been proven effective in treating hypertensive patients in China. However, the mechanisms of action of UR in reducing hypertension and myocardial fibrosis are still unclear. The purpose of this study was to explore the role of UR in an angiotensin Ⅱ (Ang Ⅱ) induced mouse model. The mice were randomly divided into 5 groups and infused with Ang Ⅱ (500 ng/kg/min) or saline, then administered UR (0.78, 1.56 or 3.12 g/kg/d) or saline for 4 weeks. UR treatment significantly attenuated the elevation of blood pressure caused by Ang Ⅱ. It enhanced myocardial function and attenuated the increase in the heart weight index and the pathological changes in the Ang Ⅱ-induced hypertensive mice. Furthermore, UR treatment inhibited cardiac fibrosis and significantly down-regulated collagen I, collagen Ⅲ, and α-SMA protein expression in cardiac tissues. UR also attenuated the expression of RhoA, ROCK1, CTGF, and TGF-ß1. In cultured cardiac fibroblasts stimulated with Ang Ⅱ, UR significantly down-regulated the expression of Collagen I, Collagen III, RhoA, ROCK1, and α-SMA. In summary, UR can significantly attenuate Ang Ⅱ-induced hypertension and cardiac fibrosis, partly via suppression of the RhoA/ROCK1 signaling pathway.

Granulocytic myeloid-derived suppressor cells increase infection risk via the IDO/IL-10 pathway in patients with hepatitis B virus-related liver failure.

Hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF) results in high susceptibility to infection. Although granulocytic myeloid-derived suppressor cells (gMDSC) are elevated in patients with HBV-ACLF, their role in HBV-ACLF pathogenesis is unknown. To elucidate the mechanism of gMDSC expansion and susceptibility to infection in HBV-ACLF patients, we analyzed the proportion of gMDSC in the peripheral blood and organ tissues of patients with HBV-ACLF and an ACLF mouse model established by continuous injection (eight times) of Concanavalin by flow cytometry and immunohistochemistry. We found that the proportion of gMDSC increased significantly in the blood and liver of patients with HBV-ACLF. This increase was positively correlated with disease severity, prognosis, and infection. gMDSC percentages were higher in peripheral blood, liver, spleen, and bone marrow than control levels in the ACLF mouse model. Immunofluorescence revealed that the gMDSC count increased in the liver of patients with HBV-ACLF as well as in the liver and spleen of ACLF mice. We further exposed peripheral blood monocyte cells from healthy donors to plasma from HBV-ACLF patients, recombinant cytokines, or their inhibitor, and found that TNF-α led to gMDSC expansion and significant upregulation of indoleamine 2, 3-dioxygenase (IDO), while blocking TNF-α signaling decreased gMDSC. Moreover, we detected proliferation and cytokine secretion of T lymphocytes when purified gMDSC was co-cultured with Pan T cells or IDO inhibitor and found that TNF-α-induced gMDSC inhibited T cell proliferation and interferon-γ production through the IDO signaling pathway. Lastly, the ability of gMDSC to phagocytose bacteria was low in patients with HBV-ACLF. Our findings elucidate HBV-ACLF pathogenesis and provide potential therapeutic targets.

Targeting NUFIP1 Suppresses Growth and Induces Senescence of Colorectal Cancer Cells.

NUFIP1 is an RNA-binding protein that interacts with fragile X mental retardation protein (FMRP) in the messenger ribonucleoprotein particle (mRNP). We previously showed that NUFIP1 was upregulated in colorectal cancer (CRC), but how the protein may contribute to the disease and patient prognosis is unknown. Here we combine database analysis, microarray, quantitative PCR, and immunohistochemistry of patients' samples to confirm our previous findings on NUFIP1 overexpression in CRC, and to reveal that increased expression of NUFIP1 in CRC tissues correlated with worse overall, recurrence-free, event-free and disease-free survival in patients, as well as with more advanced CRC clinicopathological stage. Loss of function analysis demonstrated that NUFIP1 knockdown suppressed cell growth in vitro and in vivo, inhibited cell viability and survival, and induced cell cycle arrest and apoptosis in vitro, as well as up-regulated Bax and down-regulated Bcl-2 protein expression. In addition, as a natural anticancer triterpene from various fruits and vegetables, ursolic acid (UA) treatment suppressed cell proliferation, down-regulated NUFIP1 protein expression, and further enhanced the effects of NUFIP1 knockdown in CRC cells in vitro. NUFIP1 knockdown up-regulated the expression of 136 proteins, down-regulated the expression of 41 proteins, and enriched multiple signaling pathways including the senescence-associated heterochromatin foci (SAHF) pathway. Furthermore, NUFIP1 knockdown enhanced the expression of senescence-associated-ß-galactosidase (SA-ß-gal), the SAHF markers HP1γ and trimethylation (H3k9me3), and the senescence-related protein HMGA2, as well as both p53 and its downstream p21 protein expression. Our findings suggest that NUFIP1 is overexpressed in CRC and correlates with disease progression and poor patient survival. NUFIP1 may exert oncogenic effects partly by altering senescence. UA may show potential to treat CRC by down-regulating NUFIP1.

Early Prognosis Effect of Cellular Immune Paralysis on Brain Complications of Extracorporeal Membrane Oxygenation Children with Severe Sepsis.

OBJECTIVE: The aim of the study was to explore the relationship between criticality, brain complications, and immune mechanisms in extracorporeal membrane oxygenation (ECMO) children with pneumonia and severe sepsis. METHODS: Patients with simple pneumonia (group I), ECMO patients with pneumonia and severe sepsis accompanied by brain complications (group II), and those without brain complication (group III) admitted to our pediatric intensive care unit were selected to be investigated. The relationship among the peripheral blood subgroups of immune cells, immune factors, adaptive immune responses, endothelial factors, and criticality and brain complications was then studied. RESULTS: Severe paralysis of normal immunity, excess abnormal immunity, and endothelial injury were consistent with the increase in the absolute value of base excess, lactic acid (Lac) content, and average hospitalization days and brain complications involved in group II (vs. group I). The ratio of CD63+ macrophage and CD63+ neutrophil subpopulation increased (p < 0.05); the expression levels of elastase+ neutrophil denatured subgroup (p < 0.05), the ratio of CCR2highCX3CR1low/CCR2lowCX3CR1high of macrophages and neutrophils (p < 0.0001), high-mobility group box 1 (HMGB1), YTHDF1, interleukin-17 protein and mRNA, and RAGE gene decreased to some extent (p < 0.05); the expression levels of Th1 cells chemokine CXCL9 protein and mRNA and sTIE2 protein increased to some extent (p < 0.05); the adaptive immune response of CD8+ CTL stimulated by lipopolysaccharide (LPS) was slightly enhanced (p < 0.05) in group III(vs. group II), which was consistent with the improvement of criticality, average hospitalization days, and the absence of brain complications in group III (vs. group II). CONCLUSION: ECMO support with brain complication was related to the upregulation of HMGB1 and YTHDF1 protein; the decreased number of CD63+ macrophages and neutrophils; the increased denatured neutrophil subgroup, especially the upregulated ratio of CCR2highCX3CR1low/CCR2lowCX3CR1high of macrophages and neutrophils; the imbalance of Th17/Th1, LPS-specific CD8+ CTL adaptive immune response paralysis; and the reduced endothelial sTIE2 protein expression level which caused clinical deterioration and prolonged average hospitalization days.

Qingda Granule Attenuates Angiotensin II-Induced Blood Pressure and Inhibits Ca2+/ERK Signaling Pathway.

Objective: As a well-known traditional Chinese medicine formula prescribed by academician Ke-ji Chen, Qingda granule (QDG) lowered the blood pressure of spontaneously hypertensive rats and attenuated hypertensive cardiac remodeling and inflammation. However, its functional role and underlying mechanisms on hypertensive vascular function remain largely unclear. This study aims to assess the effects of QDG treatment on Angiotensin II- (AngII-) induced hypertension and vascular function and explore its underlying mechanisms both in vitro and in vivo. Methods: In an in vivo study, 25 male C57BL/6 mice were randomly divided into five groups, including Control, AngII, AngII + QDG-L, AngII + QDG-M, and AngII + QDG-H groups (n = 5 for each group). Mice in AngII and AngII + QDG-L/-M/-H groups were infused with AngII (500 ng/kg/min), while in the Control group, they were infused with saline. Mice in AngII + QDG were intragastrically given different concentrations of QDG (0.5725, 1.145, or 2.29 g/kg/day), while in Control and AngII groups, they were intragastrically given equal volumes of double distilled water for 2 weeks. Blood pressure was determined at 0, 1, and 2 weeks of treatment. Ultrasound was used to detect the pulse wave velocity (PWV) and HE staining to detect the pathological change of the abdominal aorta. RNA sequencing (RNA-seq) was performed to identify the differentially expressed transcripts (DETs) and related signaling pathways. IHC was used to detect the expression of p-ERK in the abdominal aorta. Primary isolated rat vascular smooth muscle cells (VSMCs) were used to assess the cellular Ca2+ release and activation of the ERK pathway by confocal microscope and western blotting analysis, respectively. Results: QDG treatment significantly alleviated the elevated blood pressure, the PWV, and the thickness of the abdominal aorta in AngII-induced hypertensive mice. RNA-seq and KEGG analyses identified 1,505 DETs and multiple enriched pathways (including vascular contraction and calcium signaling pathway) after QDG treatment. Furthermore, confocal microscope showed that QDG treatment partially attenuated the increase of Ca2+ release with the stimulation of AngII in cultured VSMCs. In addition, IHC and western blotting indicated that QDG treatment also partially alleviated the increase of phospho-ERK levels in abdominal aorta tissues of mice and cultured VSMCs after the infusion or stimulation of AngII. Conclusion: QDG treatment attenuated the elevation of blood pressure, abdominal aorta dysfunction, pathological changes, Ca2+ release, and activation of the ERK signaling pathway.

Prognostic and immunological roles of Fc fragment of IgG binding protein in colorectal cancer.

Valuable diagnostic and prognostic biomarkers are urgently needed for colorectal cancer (CRC), which is one of the leading causes of mortality worldwide. Previous studies have reported altered expression of a mucin-like protein Fc fragment of IgG binding protein (FCGBP) in various types of cancer, but its potential diagnostic, prognostic and immunological roles in CRC remain to be determined. Therefore, the aim of current study was to investigate the potential roles of FCGBP in CRC. The present study investigated FCGBP mutations and changes in its expression levels using a combination of microarray and public dataset analyses, as well as immunohistochemistry. The results demonstrated a 10.5% mutation frequency in the FCGBP coding sequence in CRC tissues, and identified decreased FCGBP mRNA or protein expression levels in colorectal adenoma and CRC (compared with those in normal colorectal tissues from healthy control subjects), including pathologically advanced CRC (stage III+IV vs. I+II). Survival analysis using the GEPIA and Kaplan-Meier Plotter databases revealed that low FCGBP expression levels were associated with short overall, disease-free, relapse-free and event-free survival times in patients with CRC. Notably, analysis using the online Tumor IMmune Estimation Resource database revealed a positive correlation between FCGBP expression levels and the extent of infiltrating immune cells, such as B cells and dendritic cells. Consistently, the expression levels of most markers (51/57) for various types of immune cells were significantly correlated with FCGBP expression levels in CRC tissues. These findings suggested that FCGBP may serve as a diagnostic and prognostic biomarker, and that FCGBP may be associated with immune infiltration in CRC.

Huoxin Pill inhibits isoproterenol-induced transdifferentiation and collagen synthesis in cardiac fibroblasts through the TGF-ß/Smads pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The abnormal proliferation and differentiation of cardiac fibroblasts (CFs) are universally regarded as the key process for the progressive development of cardiac fibrosis following various cardiovascular diseases. Huoxin Pill (Concentrated pill, HXP) is a Chinese herbal formula for treating coronary heart disease. However, the cellular and molecular mechanisms of HXP in the treatment of myocardial fibrosis are still unclear. AIM OF THE STUDY: To investigate the effects of HXP on CFs transdifferentiation and collagen synthesis under isoproterenol (ISO) conditions, as well as the potential mechanism of action. MATERIALS AND METHODS: In vivo, we established a rat model of cardiac fibrosis induced by ISO, and administered with low or high dose of HXP (10 mg/kg/day or 30 mg/kg/day). The level of α-SMA was detected by immunohistochemistry examination, and combined with RNA-sequencing analysis to determine the protective effect of HXP on myocardial fibrosis rats. In vitro, by culturing primary rat CFs, we examined the effects of HXP on the proliferation and transdifferentiation of CFs using CCK8, scratch wound healing and immunofluorescence assays. Western blot was used to determine protein expression. RESULTS: The findings revealed that HXP protects against ISO-induced cardiac fibrosis and CFs transdifferentiation in rats. RNA-sequencing and pathway analyses demonstrated 238 or 295 differentially expressed genes (DEGs) and multiple enriched signal pathways, including transforming growth factor-beta (TGF-ß) receptor signaling activates Smads, downregulation of TGF-ß receptor signaling, signaling by TGF-ß receptor complex, and collagen formation under treatment with low or high-dose of HXP. Moreover, HXP also markedly inhibited ISO-induced primary rat CFs proliferation, transdifferentiation, collagen synthesis and the upregulation of TGF-ß1 and phosphorylated Smad2/3 protein expression. CONCLUSION: HXP suppresses ISO-induced CFs transdifferentiation and collagen synthesis, and it may exert these effects in part by inhibiting the activation of the TGF-ß/Smads pathway. This may be a new therapeutic tool for cardiac fibrosis.

Qingda granule attenuates cardiac fibrosis via suppression of the TGF-ß1/Smad2/3 signaling pathway in vitro and in vivo.

Cardiac fibrosis plays an important role in hypertension-related contractile dysfunction and heart failure. Qingda granule (QDG), derived from the Qingxuan Jiangya decoction, has been used clinically for more than 60 years to treat hypertension. However, the effect of QDG on hypertensive cardiac fibrosis remains largely unknown. The objective of this study was to investigate the effect of QDG on cardiac fibrosis and explore the underlying mechanism in vivo and in vitro. For in vivo experiments, 30 male spontaneously hypertensive rats were randomly divided into groups that received no QDG or one of three doses (0.45, 0.9 or 1.8 g/kg/day). Positive-control animals received valsartan (VAL, 7.2 mg/kg/day). Treatments were administered by gavage for 10 weeks. All three doses of QDG and VAL led to significantly lower blood pressure than in SHR animals. Besides, all three doses of QDG and VAL attenuated pathological changes in SHR animals. However, only intermediate, high concentrations of QDG and VAL led to significantly lower left ventricle ejection fraction and left ventricle fractional shortening than in SHR animals. Therefore, the minimum and effective QDG dose (intermediate concentration of QDG) was selected for subsequent animal experiments in this study. Our results showed that intermediate concentration of QDG also significantly mitigated the increases in levels of α-smooth muscle actin (α-SMA), proliferating cell nuclear antigen (PCNA), collagen III, transforming growth factor-ß1 (TGF-ß1) and in the ratio of phospho-Smad2/3 to total Smad2/3 protein in cardiac tissue, based on immunohistochemistry, Western blotting, and Masson staining. For in vitro experiments, primary cardiac fibroblasts were stimulated with 100 nM angiotensin II in the presence or absence of QDG. And we tested different concentrations of QDG (3.125, 6.25, 12.5, 25, 50 µg/mL) in the cell viability experiment. Our results showed that 3.125, 6.25 and 12.5 µg/mL of QDG treatment for 24 h didn't affect the cell viability of cardiac fibroblasts. Consistently, QDG at 6.25 or 12.5 µg/mL significantly reduced cell viability and down-regulated α-SMA in primary cardiac fibroblasts were stimulated with 100 nM angiotensin II. Therefore, QDG at 12.5 µg/mL was chosen for the following cell experiment. Our results showed that QDG at 12.5 µg/mL alleviated the increase of PCNA, collagen Ⅲ, TGF-ß1 expression, and the ratio of phospho-Smad2/3 to total Smad2/3 protein. Our studies in vitro and in vivo suggest that QDG reduces blood pressure and cardiac fibrosis as well as protecting cardiac function, and that it exerts these effects in part by suppressing TGF-ß1/Smad2/3 signaling.

Huoxin Pill () Attenuates Cardiac Fibrosis by Suppressing TGF-ß1/Smad2/3 Pathway in Isoproterenol-Induced Heart Failure Rats.

OBJECTIVE: To evaluate the effects of Huoxin Pill (, HXP) on cardiac fibrosis and heart failure (HF) in isoproterenol (ISO)-induced HF rats. METHODS: Thirty Wistar rats were randomly divided into 5 groups including control, HF, isosorbide mononitrate (ISMN), HXP low (HXP-L), and HXP high (HXP-H) groups (n=6 for each group) according to the complete randomization method. Rats were pretreated with ISMN (5 mg/kg daily), low concentration of HXP (10 mg/kg daily) or high concentration of HXP (30 mg/kg daily) or equal volume of saline by intragastric administration for 1 week, followed by intraperitoneal injection of ISO (10 mg/kg, 14 days), and continually intragastric administrated with above medicines or saline for additional 6 weeks. The effects of HXP treatment on the cardiac function, heart weight index (HWI), pathological changes, and collagen content were further assessed. Moreover, the role of HXP on activation of transforming growth factor- ß 1 (TGF-ß 1)/Smads pathway was further explored using immunohistochemistry (IHC) and Western-blot assay. RESULTS: HXP treatment significantly alleviated the decrease of ejection fraction (EF) and fractional shortening (FS), while decreased the elevation of left ventricular end-systolic volume (LVESV) in ISO-induced HF rats (P<0.05). Moreover, HXP treatment obviously attenuated the increase of HWI and serum level of creatine kinase MB (CK-MB, P<0.05), as well as pathological changes in ISO-induced HF rats. Further determination indicated that HXP treatment alleviated the elevation of collagen I and collagen III protein expression in cardiac tissues of ISO-induced HF rats. Furthermore, HXP treatment significantly down-regulated the increase of TGF-ß 1 and p-Smad2/3 protein expression in cardiac tissues of HF rats (P<0.05), while did not affect the expression of total Smad2/3. CONCLUSIONS: HXP attenuated heart failure and cardiac fibrosis in ISO-induced HF rats by suppression of TGF-ß 1/Smad2/3 pathway.

Qingda granule attenuates angiotensin II-induced cardiac hypertrophy and apoptosis and modulates the PI3K/AKT pathway.

Qingda granule (QDG), simplified from Qingxuan Jiangya Decoction, is a well-known traditional Chinese medicine formula that has been used for decades to treat hypertension. However, the cardioprotective effects of QDG on Ang II-induced hypertension remain unknown. This study aimed to investigate the effects of QDG on hypertension-induced cardiac hypertrophy and apoptosis, as well as explore its underlying mechanisms. Mice were infused with Ang II (500 ng/kg/min) or saline solution as control, then administered oral QDG (1.145 g/kg/day) or saline for two weeks. QDG treatment attenuated the elevation in blood pressure caused by Ang II, as well as the decreased left ventricle ejection fractions and fractional shortening. Moreover, QDG treatment significantly alleviated the Ang II-induced elevation of the ratio of heart weight to tibia length, as well as cardiac injury, hypertrophy, and apoptosis. In cultured H9C2 cells stimulated with Ang II, QDG partially reversed the increase in cell surface area and number of apoptotic cells, up-regulation of hypertrophy markers ANP and BNP, and activation of caspases-9 and -3. QDG also partially reversed Ang II-induced accumulation of reactive oxygen species (ROS), depolarization of the mitochondrial membrane, release of cytochrome C, up-regulation of Bax, and decrease in levels of p-PI3K, p-AKT, and Bcl-2. These results suggest that QDG can significantly attenuate Ang II-induced hypertension, cardiac hypertrophy and apoptosis, and it may exert these effects in part by suppressing ROS production and activating the PI3K/AKT signaling pathway.

Down-Regulating HAUS6 Suppresses Cell Proliferation by Activating the p53/p21 Pathway in Colorectal Cancer.

Background: HAUS6 participates in microtubule-dependent microtubule amplification, but its role in malignancies including colorectal cancer (CRC) has not been explored. We therefore assessed the potential oncogenic activities of HAUS6 in CRC. Results: HAUS6 mRNA and protein expression is higher in CRC tissues, and high HAUS6 expression is correlated with shorter overall survival in CRC patients. HAUS6 knockdown in CRC cell lines suppressed cell growth in vitro and in vivo by inhibiting cell viability, survival and arresting cell cycle progression at G0/G1, while HAUS6 over-expression increased cell viability. We showed that these effects are dependent on activation of the p53/p21 signalling pathway by reducing p53 and p21 degradation. Moreover, combination of HAUS6 knockdown and 5-FU treatment further enhanced the suppression of cell proliferation of CRC cells by increasing activation of the p53/p21 pathway. Conclusion: Our study highlights a potential oncogenic role for HAUS6 in CRC. Targeting HAUS6 may be a promising novel prognostic marker and chemotherapeutic target for treating CRC patients.

Transcription Factor EBF1 Over-Expression Suppresses Tumor Growth in vivo and in vitro via Modulation of the PNO1/p53 Pathway in Colorectal Cancer.

Early B cell factor 1 (EBF1) has been identified as an upstream transcription factor of the potential oncogene PNO1 and is involved in the growth of colorectal cancer (CRC) cells. However, its expression, biological function, and underlying mechanism of action in most solid tumors remain largely unknown. We postulated that EBF1 has a role in the pathophysiology of CRC. Analysis of EBF1 mRNA expression in CRC tumor samples from several public databases and directly from banked tissues revealed that EBF1 mRNA expression is lower in CRC tissue compared to non-cancerous colorectal tissue. Survival analysis of multiple datasets revealed that low EBF1 expression was correlated with shorter overall survival, relapse-free survival, and event-free survival in CRC patients. Transduction of lentivirus encoding full length EBF1 followed by in vitro and in vivo assays demonstrated that EBF1 over-expression in CRC cell lines suppresses cell growth by inhibiting cell viability, cell survival, and induces cell cycle arrest and apoptosis. Mechanistic investigation indicated that EBF1 over-expression down-regulates PNO1 mRNA and protein expression, as well as transcriptional activity while up-regulating the expression of p53 and p21 proteins. These findings suggest that EBF1 is a novel potential tumor suppressor in CRC with prognostic value for the identification of patients at high-risk of relapse.

Huoxin pill attenuates myocardial infarction-induced apoptosis and fibrosis via suppression of p53 and TGF-ß1/Smad2/3 pathways.

Huoxin Pill (HXP), a Traditional Chinese Medicine, is used widely to treat patients with coronary heart disease and angina pectoris in China. However, the underlying protective mechanism of HXP on cardiac apoptosis and fibrosis has never been evaluated. Therefore, the aim of this study was to investigate the role of HXP in a myocardial infarction (MI) mouse model. The mice were randomly divided into 3 groups and subjected to surgical ligation of the left anterior descending (LAD) coronary artery or sham surgery (n = 6 for each group) and treated with HXP (50 mg/kg/day) or saline by gavage for 2 weeks. At 2 weeks post MI, we found that HXP significantly enhanced myocardial function and attenuated the increase of heart weight index (HWI) and pathological changes in MI mice. RNA-sequencing and KEGG pathway analyses identified 660 differentially expressed genes and multiple enriched signaling pathways including p53 and TGF-ß. In support of these findings, HXP attenuated cardiac apoptosis and decreased p53 and Bax protein expression, while increasing Bcl-2 protein expression in cardiac tissues of MI mice. Furthermore, HXP treatment inhibited cardiac fibrosis and significantly down-regulated TGF-ß1 protein expression and Smad2/3 phosphorylation in cardiac tissues. In summary, HXP can improve cardiac function in mice after MI by attenuating cardiac apoptosis and fibrosis partly via supression of the p53/Bax/Bcl-2 and TGF-ß1/Smad2/3 pathways.

Antihypertensive and Vasodilatory Effects of Qingda Granules by Suppression of Calcium Influx and the AKT Pathway.

The Qingda granule (QDG) formulation was simplified from the Qingxuan Jiangya Decoction, which has been used in China to treat hypertension for decades. However, the molecular mechanisms of QDG in antihypertension remain largely unknown. Therefore, we evaluated the therapeutic efficacy of QDG against elevated blood pressure and explored its underlying mechanism. QDG treatment decreased elevated blood pressure and increased the vascular elasticity of thoracic aortic rings to KCl stimulation in spontaneously hypertensive rats. QDG treatment increased the relaxation of isolated thoracic aortic rings precontracted with norepinephrine (NE) or KCl in an endothelium-independent manner, which was attenuated by treatment with verapamil, but not by treatment with TEA, 4-AP, Gli, or BaCl2. Moreover, QDG pretreatment attenuated the CaCl2-induced constriction of isolated thoracic aortic rings in K- or NE-containing Ca-free solutions. In addition, QDG pretreatment significantly inhibited the influx of Ca in A7r5 cells induced by a K- or NE-containing Ca solution and decreased the levels of p-AKT but had no effect on levels of total AKT protein in isolated thoracic aortic rings. Considering these results, QDG treatment attenuated elevated blood pressure and promoted the vasorelaxation of thoracic aortic rings by inhibiting the influx of Ca and activating the AKT pathway.

Development and Validation of a Novel Model to Predict Liver Histopathology in Patients with Chronic Hepatitis B.

It is still vague for chronic hepatitis B (CHB) patients with normal or mildly increasing alanine aminotransferase (ALT) level to undergo antiviral treatment or not. The purpose of our study was to establish a noninvasive model based on routine blood test to predict liver histopathology for antiviral therapy. This retrospective study enrolled 258 CHB patients with liver biopsy from the First Hospital of Quanzhou (training cohort, n=126) and Huashan Hospital (validation cohort, n=132). Histologic grading of necroinflammation (G) and liver fibrosis (S) was performed according to the Scheuer scoring system. A novel model, ATPI, including aspartate aminotransferase (AST), total bilirubin (TBil), and platelets (PLT), was developed in training cohort. The area under ROC curves (AUC) of ATPI for predicting antiviral therapy indication was 0.83 in training cohort and was 0.88 in the validation cohort, respectively. Similarly, ATPI also displayed the highest AUC in predicting antiviral therapy indication in CHB patients with normal or mildly increasing ALT level. In conclusion, ATPI is a novel independent model to predict liver histopathology for antiviral therapy in CHB patients with normal and mildly increased ALT levels.

EBF1-Mediated Upregulation of Ribosome Assembly Factor PNO1 Contributes to Cancer Progression by Negatively Regulating the p53 Signaling Pathway.

The RNA-binding protein PNO1 is critical for ribosome biogenesis, but its potential role in cancer remains unknown. In this study, online data mining, cDNA, and tissue microarrays indicated that PNO1 expression was higher in colorectal cancer tissue than in noncancerous tissue, and its overexpression was associated with worse patient survival. Gain-of-function and loss-of-function studies demonstrated that PNO1 knockdown suppressed growth of colorectal cancer cells in vitro and in vivo, while PNO1 overexpression promoted colorectal cancer cell proliferation in vitro. In colorectal cancer cells expressing wild-type p53, PNO1 knockdown enhanced expression of p53 and its downstream gene p21, and reduced cell viability; these effects were prevented by p53 knockout and attenuated by the p53 inhibitor PFT-α. Moreover, PNO1 knockdown in HCT116 cells decreased levels of 18S rRNA, of 40S and 60S ribosomal subunits, and of the 80S ribosome. It also reduced global protein synthesis, increasing nuclear stress and inhibiting MDM2-mediated ubiquitination and p53 degradation. Overexpressing EBF1 suppressed PNO1 promoter activity and decreased PNO1 mRNA and protein, inhibiting cell proliferation and inducing cell apoptosis through the p53/p21 pathway. In colorectal cancer tissues, the expression of EBF1 correlated inversely with PNO1. Data mining of online breast and lung cancer databases showed increased PNO1 expression and association with poor patient survival; PNO1 knockdown reduced cell viability of cultured breast and lung cancer cells. Taken together, these findings indicate that PNO1 is overexpressed in colorectal cancer and correlates with poor patient survival, and that PNO1 exerts oncogenic effects, at least, in part, by altering ribosome biogenesis. SIGNIFICANCE: This study identifies the ribosome assembly factor PNO1 as a potential oncogene involved in tumor growth and progression of colorectal cancer.