Osthole prevents tamoxifen-induced liver injury in mice.

Tamoxifen (TMX) is an antiestrogen drug that is used in the treatment and prevention of all stages of estrogen-dependent breast cancer. Adverse effects of TMX include hepatotoxicity. In this study, we investigated the therapeutic effects of osthole, isolated from medicinal plants especially Fructus Cnidii, on TMX-induced acute liver injury in mice. Mice were injected with osthole (100 mg/kg, ip) or vehicle, followed by TMX (90 mg/kg, ip) 24 h later. We showed that a single injection of TMX-induced liver injury and oxidative stress. Pretreatment with osthole attenuated TMX-induced liver injury evidenced by dose-dependent reduction of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities. Pretreatment with osthole also blunted TMX-induced oxidative stress, evidenced by significant increase of reduced glutathione (GSH) as well as reduction of malondialdehyde (MDA) and hydrogen peroxide (H2O2). Consistently, osthole significantly enhanced the expressions of antioxidant genes (GPX1, SOD2, GCL-c, and G6pdh), but suppressed those of pro-oxidant genes (NOX2 and ACOX). Furthermore, osthole inhibited the production of inflammatory cytokines, reduced the metabolic activation of TMX, and promoted its clearance. We further revealed that osthole elevated hepatic cAMP and cGMP levels, but inhibition of PKA or PKG failed to abolish the hepatoprotective effect of osthole. Meanwhile, prominent phosphorylation of p38 was observed in liver in response to TMX, which was significantly inhibited by osthole. Pretreatment with SB203580, a p38 inhibitor, significantly attenuated TMX-induced increase of ALT and AST activities, reduced oxidative stress, and reversed the alterations of gene expression caused by TMX. Moreover, pretreatment with L-buthionine sulfoximine (BSO), an inhibitor of GSH synthesis, partly reversed the effect of osthole on TMX-induced liver injury. Consistently, pretreatment with N-acetyl-L-cysteine (NAC) significantly attenuated TMX-induced increase in ALT and AST activities. Notably, both BSO and NAC had no detectable effect on the phosphorylation levels of p38. Collectively, our results suggest that osthole prevents TMX hepatotoxicity by suppressing p38 activation and subsequently reducing TMX-induced oxidative damage.

Four-year variation in pathogen distribution and antimicrobial susceptibility of urosepsis: a single-center retrospective analysis.

Background: Urosepsis is a common disease in urology, which is characterized by high treatment costs and high mortality. In the treatment of sepsis, anti-infection therapy is the most important means. However, the effect of empirical anti-infection therapy is often not ideal. Therefore, it is necessary to continuously monitor the prevalence of bacterial isolates in the blood culture of patients with urinary sepsis and their sensitivity to antibacterial drugs. This is of great significance to improve the efficacy of empirical antibiotic therapy for urosepsis. Objective: To elucidate the landscape of prevailing bacterial profiles and their antimicrobial susceptibilities in urosepsis cases, and to furnish robust clinical evidence to underpin the timely initiation of empirical antibiotic treatment. Methods: Collect the basic information and blood culture results of patients with urosepsis hospitalized from 2017 to 2020. Retrospective analysis of bacterial species and antimicrobial susceptibility in urosepsis and changes over 4 years. Results: Gram-negative bacteria (178 isolates, 75.11%) constituted the main pathogens causing urosepsis, followed by Gram-positive bacteria (46 isolates, 19.41%) and fungus (13 isolates, 5.48%). The sensitivity of ertapenem, meropenem, amikacin, and imipenem to Gram-negative bacteria all exceeded 85%. The sensitivity rates of levofloxacin, gentamicin, and ciprofloxacin are decreasing every year (p < 0.05). Tigecycline, vancomycin, and linezolid exhibited excellent sensitivity against Gram-positive bacteria. Among fungi, fluconazole demonstrated universal sensitivity, while itraconazole-resistant isolates have been found, and amphotericin B is still effective. Conclusion: Analysis of blood culture results of patients more accurately reflected the etiology of urosepsis, mainly Escherichia coli, Enterococcus, and Klebsiella pneumoniae. If there are no definitive blood culture results, empiric treatment of urosepsis should not include fluoroquinolone antibiotics. Cefepime, cefoxitin, and ceftazidime are the most sensitive antibiotics to Gram-negative bacteria besides carbapenem antibiotics. In addition, the current situation regarding extended-spectrum ß-lactamase-producing bacteria and carbapenem-resistant Enterobacteriaceae bacteria resistance is extremely concerning with limited therapeutic options available. Strengthening antibiotic management practices and exploring novel antibacterial agents can help mitigate this issue.

Unbiased proteomic analysis detects painful systemic inflammatory profile in the serum of nerve-injured mice.

ABSTRACT: Neuropathic pain is a complex, debilitating disease that results from injury to the somatosensory nervous system. The presence of systemic chronic inflammation has been observed in patients with chronic pain but whether it plays a causative role remains unclear. This study aims to determine the perturbation of systemic homeostasis by an injury to peripheral nerve and its involvement in neuropathic pain. We assessed the proteomic profile in the serum of mice at 1 day and 1 month after partial sciatic nerve injury (PSNL) or sham surgery. We also assessed mouse mechanical and cold sensitivity in naïve mice after receiving intravenous administration of serum from PSNL or sham mice. Mass spectrometry-based proteomic analysis revealed that PSNL resulted in a long-lasting alteration of serum proteome, where most of the differentially expressed proteins were in inflammation-related pathways, involving cytokines and chemokines, autoantibodies, and complement factors. Although transferring sham serum to naïve mice did not change their pain sensitivity, PSNL serum significantly lowered mechanical thresholds and induced cold hypersensitivity in naïve mice. With broad anti-inflammatory properties, bone marrow cell extracts not only partially restored serum proteomic homeostasis but also significantly ameliorated PSNL-induced mechanical allodynia, and serum from bone marrow cell extracts-treated PSNL mice no longer induced hypersensitivity in naïve mice. These findings clearly demonstrate that nerve injury has a long-lasting impact on systemic homeostasis, and nerve injury-associated systemic inflammation contributes to the development of neuropathic pain.

Population pharmacokinetic analysis, renal safety, and dosing optimization of polymyxin B in lung transplant recipients with pneumonia: A prospective study.

Objectives: This study aims to characterize the population pharmacokinetics of polymyxin B in lung transplant recipients and optimize its dosage regimens. Patients and methods: This prospective study involved carbapenem-resistant organisms-infected patients treated with polymyxin B. The population pharmacokinetic model was developed using the NONMEM program. The clinical outcomes including clinical treatment efficacy, microbiological efficacy, nephrotoxicity, and hyperpigmentation were assessed. Monte Carlo simulation was performed to calculate the probability of target attainment in patients with normal or decreased renal function. Results: A total of 34 hospitalized adult patients were included. 29 (85.29%) patients were considered of clinical cure or improvement; 14 (41.18%) patients had successful bacteria elimination at the end of the treatment. Meanwhile, 5 (14.71%) patients developed polymyxin B-induced nephrotoxicity; 19 (55.88%) patients developed skin hyperpigmentation. A total of 164 concentrations with a range of 0.56-11.66 mg/L were obtained for pharmacokinetic modeling. The pharmacokinetic characteristic of polymyxin B was well described by a 1-compartment model with linear elimination, and only creatinine clearance was identified as a covariate on the clearance of polymyxin B. Monte Carlo simulations indicated an adjusted dosage regimen might be needed in patients with renal insufficiency and the currently recommended dose regimens by the label sheet of polymyxin B may likely generate a subtherapeutic exposure for MIC = 2 mg/L. Conclusion: Renal function has a significant effect on the clearance of polymyxin B in lung transplant recipients, and an adjustment of dosage was needed in patients with renal impairments.

Does Low Grade Systemic Inflammation Have a Role in Chronic Pain?

One of the major clinical manifestations of peripheral neuropathy, either resulting from trauma or diseases, is chronic pain. While it significantly impacts patients' quality of life, the underlying mechanisms remain elusive, and treatment is not satisfactory. Systemic chronic inflammation (SCI) that we are referring to in this perspective is a state of low-grade, persistent, non-infective inflammation, being found in many physiological and pathological conditions. Distinct from acute inflammation, which is a protective process fighting against intruders, SCI might have harmful effects. It has been associated with many chronic non-communicable diseases. We hypothesize that SCI could be a predisposing and/or precipitating factor in the development of chronic pain, as well as associated comorbidities. We reviewed evidence from human clinical studies indicating the coexistence of SCI with various types of chronic pain. We also collated existing data about the sources of SCI and who could have it, showing that those individuals or patients having SCI usually have higher prevalence of chronic pain and psychological comorbidities. We thus elaborate on the need for further research in the connection between SCI and chronic pain. Several hypotheses have been proposed to explain these complex interactions.

Circulating exosomal microRNAs as novel potential detection biomarkers in pancreatic cancer.

Circulating exosomal microRNAs (ex-miRNAs) are reflective of the characteristics of the tumor and are valuable biomarkers in different types of tumor. In addition, miRNAs serve important roles in tumor progression and metastasis. The present study aimed to investigate the circulating ex-miRNA-21 and miRNA-210 as novel biomarkers for patients with pancreatic cancer (PC). For this purpose, serum ex-miRNAs were extracted from the serum of patients with PC (n=30) and chronic pancreatitis (CP) (n=10) using an RNA isolation kit. For exosome identification in serum, transmission electron micrographs were used to determine crystalline structure, western blotting was used to identify exosomal markers, and NanoSight was used for nanoparticle characterization. The relative expression levels of ex-miRNAs were quantified using quantitative PCR and compared between patients with PC and CP. The expression levels of both ex-miRNA-21 and miRNA-210 were significantly higher in patients with PC compared with patients with CP (both P<0.001). However, no significant difference in the relative serum levels of free miR-21 and miR-210 was observed between the 2 groups of patients (both P>0.05). ex-miRNA-21 and miRNA-210 were associated with tumor stage, as well as other factors. The diagnostic potential of ex-miRNA-21 and miRNA-210 levels was 83 and 85%, respectively. In addition, when ex-miRNA and serum carbohydrate antigen 19-9 expression levels were combined, the accuracy increased to 90%. The present study identified that serum ex-miRNAs, miRNA-21 and miRNA-210 may be of value as potential biomarkers and therapeutic targets for the diagnosis and treatment of PC.

[Analysis on the expression of the stem cells related genes in hepatocellular carcinoma cell lines].

OBJECTIVE: To investigate the expression of Oct4, Sox2, Nanog, SMO, beta-Catenin and Wnt5b mRNA in four hepatocellular carcinoma cell lines of SMMC-7721, Bel-7402, HepG2, MHCC-97 and normal hepatocellular cell line of L02, and to compare the response of these cell lines to all-trans retinoic acid. METHODS: RT-PCR was used to detect expression of Oct4, Sox2, Nanog, SMO, beta-Catenin and Wnt5b mRNA in four hepatocellular carcinoma cell lines and normal hepatocellular cell line. Real time-PCR was used to quantify the expression of the genes. RESULTS: There are different levels of expression of the stem cell-related gene in hepatocellular carcinoma cell lines and control cell line (P less than 0.05). There are significant differences in HepG2 and L-02 for the response to all-trans retinoic acid (P less than 0.05). CONCLUSIONS: The stem cell-related genes are differentially expressed in different hepatocellular carcinoma cell lines.

Luteolin induces hippocampal neurogenesis in the Ts65Dn mouse model of Down syndrome.

Studies have shown that the natural flavonoid luteolin has neurotrophic activity. In this study, we investigated the effect of luteolin in a mouse model of Down syndrome. Ts65Dn mice, which are frequently used as a model of Down syndrome, were intraperitoneally injected with 10 mg/kg luteolin for 4 consecutive weeks starting at 12 weeks of age. The Morris water maze test was used to evaluate learning and memory abilities, and the novel object recognition test was used to assess recognition memory. Immunohistochemistry was performed for the neural stem cell marker nestin, the astrocyte marker glial fibrillary acidic protein, the immature neuron marker DCX, the mature neuron marker NeuN, and the cell proliferation marker Ki67 in the hippocampal dentate gyrus. Nissl staining was used to observe changes in morphology and to quantify cells in the dentate gyrus. Western blot assay was used to analyze the protein levels of brain-derived neurotrophic factor (BDNF) and phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) in the hippocampus. Luteolin improved learning and memory abilities as well as novel object recognition ability, and enhanced the proliferation of neurons in the hippocampal dentate gyrus. Furthermore, luteolin increased expression of nestin and glial fibrillary acidic protein, increased the number of DCX+ neurons in the granular layer and NeuN+ neurons in the subgranular region of the dentate gyrus, and increased the protein levels of BDNF and p-ERK1/2 in the hippocampus. Our findings show that luteolin improves behavioral performance and promotes hippocampal neurogenesis in Ts65Dn mice. Moreover, these effects might be associated with the activation of the BDNF/ERK1/2 pathway.

Gene expression alterations of human liver cancer cells following borax exposure.

Borax is a boron compound that is becoming widely recognized for its biological effects, including lipid peroxidation, cytotoxicity, genotoxicity, antioxidant activity and potential therapeutic benefits. However, it remains unknown whether exposure of human liver cancer (HepG2) cells to borax affects the gene expression of these cells. HepG2 cells were treated with 4 mM borax for either 2 or 24 h. Gene expression analysis was performed using Affymetrix GeneChip Human Gene 2.0 ST Arrays, which was followed by gene ontology analysis and pathway analysis. The clustering result was validated using reverse transcription­quantitative polymerase chain reaction. A cell proliferation assay was performed using Celigo Image Cytometer Instrumentation. Following this, 2­ or 24­h exposure to borax significantly altered the expression level of a number of genes in HepG2 cells, specifically 530 genes (384 upregulated and 146 downregulated) or 1,763 genes (1,044 upregulated and 719 downregulated) compared with the control group, respectively (≥2­fold; P<0.05). Twenty downregulated genes were abundantly expressed in HepG2 cells under normal conditions. Furthermore, the growth of HepG2 cells was inhibited through the downregulation of PRUNE1, NBPF1, PPcaspase­1, UPF2 and MBTPS1 (≥1.5­fold, P<0.05). The dysregulated genes potentially serve important roles in various biological processes, including the inflammation response, stress response, cellular growth, proliferation, apoptosis and tumorigenesis/oncolysis.

Overexpression and correlation of HIF-2α, VEGFA and EphA2 in residual hepatocellular carcinoma following high-intensity focused ultrasound treatment: Implications for tumor recurrence and progression.

Rapid growth of residual tumors can occur as a result of their recurrence and progression. The present study aimed to investigate the expression of hypoxia inducible factor-2 subunit α (HIF-2α), vascular endothelial growth factor A (VEGFA), erythropoietin-producing hepatocellular A2 (EphA2) and angiogenesis in residual hepatocellular carcinoma (HCC), following treatment with high-intensity focused ultrasound (HIFU) ablation, in order to investigate the association between protein expression and tumor recurrence and growth. Athymic BALB/c (nu/nu) mice were subcutaneously inoculated with the HCC cell line HepG2, in order to create xenograft tumors. Approximately 30 days post-inoculation, eight mice were treated with HIFU, whereas eight mice received no treatment and acted as the control group. Residual tumor tissues were obtained from the experimental groups after one month. Levels of HIF-2α, VEGFA, EphA2 and cluster of differentiation 31 (CD31) expression was measured by immunohistochemical staining. CD31-positive vascular endothelial cells were counted to calculate microvascular density (MVD), and western blot analysis was performed to determine levels of HIF-2α, VEGFA, and EphA2 protein. It was found that the expression levels of HIF-2α, VEGFA, EphA2, and MVD proteins in residual HCC tissues were significantly higher than in the control group tissues (P<0.05). Tumor MVD was strongly correlated with VEGFA (R=0.957, P<0.01) and EphA2 (R=0.993, P<0.01) protein expression levels. Furthermore, there was a significant positive correlation between HIF-2α and EphA2 expression (R=0.991, P<0.01). The correlation between VEGFA and EphA2 expression was also positive (R=0.985, P<0.01). These data suggest that overexpression of HIF-2α, VEGFA and EphA2 is related to angiogenesis in residual HCC following HIFU ablation, potentially via their association with key mediators of recurrence.

Effect of CD16a, the surface receptor of Kupffer cells, on the growth of hepatocellular carcinoma cells.

FcγRIIIa (CD16) is a low-affinity Fc receptor of IgG. As the idio-binding receptor of IgG Fc, it plays an important role in the antibody-dependent cellular cytotoxicity of natural killer cells. The aim of the present study was to investigate the distribution of Kupffer cells (KCs) and the expression of their surface receptor FcγRIIIa in hepatocellular carcinoma. Furthermore, we also aimed to observe the functional mechanism of FcγRIIIa. Immunohistochemical analysis was employed to study KCs and FcγRIIIa. In order to explore the role of FcγRIIIa in the growth of cancer cells, KCs and H22 tumor cells were co-cultured in different serum. The mRNA expression levels of tumor necrosis factor (TNF)-α and FcγRIIIa were analyzed by RT-qPCR; the TNF-α and FcγRIIIa protein expression levels were examined by enzyme­linked immunosorbent assay and western blot analysis, respectively. Our results showed that the number of Kuppfer cells in cancerous tissues (21.6±7.8) was lower than those in para-cancerous (68.8±9.1) tissues and adjacent normal hepatic tissues (62.0±1.9) (P<0.01); this decreased with the reduction in the differentiation degree of cancer (P<0.05). FcγRIIIa-positive cells were similar in morphology to KCs, and their distributive tendency was coincident (P<0.05). The increase in CD16a mRNA levels in the group treated with immune serum was 3.9-, 4.9- and 3.9-fold greater than that in the ordinary serum group at different time points, and CD16a protein expression also markedly increased (P<0.05). However, these effects were inhibited by the addition of anti-IgG Fc serum (P<0.05). The results of the present study suggested that FcγRIIIa resided in KCs, and it contributed to the inhibition of the growth of liver tumor cells.

Connexin32­mediated antitumor effects of suicide gene therapy against hepatocellular carcinoma: In vitro and in vivo anticancer activity.

Normal hepatocytes express connexin32 (Cx32), which forms gap junctions at cell­cell contact areas. The aim of the present study was to investigate whether Cx32 mediates the cell death­inducing effects of ultrasound microbubbles carrying the herpes simplex virus thymidine kinase (HSV­TK) suicide gene against hepatocellular carcinoma cells in vitro and in vivo. HepG2 cells were exposed to different concentrations of trans­retinoic acid (ATRA) in culture, to evaluate the intrinsic antitumor effect of ATRA. Detailed in­vitro and in­vivo investigations on the antitumor effects of ATRA via Cx32 mediation were performed, and the possible underlying mechanisms of action of the compound were then examined. The gene expression of HSV­TK transfected by ultrasound wave irradiation in the HepG2 cells was quantified using reverse transcription­quantitative polymerase chain reaction analysis. The effects on cell death were assessed using an MTT assay. The protein expression levels of Cx32 in ATRA­untreated or ATRA­treated tissues were quantified by immunohistochemical analysis and Western blot assays. The HSV­TK gene was successfully transfected into the HepG2 cell using ultrasound wave irradiation, and was stably expressed. Compared with the other groups, the HSV­TK gene group treated with ATRA exhibited an increased number of apoptotic cells (P<0.05) and improved tumor suppression (P<0.05). ATRA significantly increased the expression of Cx32 in the hepatoma tissues (P<0.01). The present study demonstrated that ATRA elevated the protein expression of Cx32 and enhanced the bystander effect of the HSV­TK/GCV suicide gene therapy system, which may provide a potential strategy for hepatocellular carcinoma treatment.

Assessing apoptosis gene expression profiling with a PCR array in the hippocampus of Ts65Dn mice.

It is well known that Down syndrome (DS) is a condition in which extra genetic material causes delays in the way a child develops, both mentally and physically. Intellectual disability is the foremost and most debilitating trait, which caused loss of cognitive abilities and the development of early onset Alzheimer's disease (AD). Ts65Dn mice were used in this study. We isolated the hippocampus. First, we used transmission scanning electron microscopy to directly observe the hippocampus and confirm if apoptosis had occurred. Second, we customized a PCR array with 53 genes, including several important genes related to cell apoptosis. Gene expression was detected by RT-PCR. There were varying degrees of changes characteristic of apoptosis in the hippocampus of Ts65Dn mice, which mainly included the following: nuclear membrane thinning, unevenly distributed chromosomes, the production of chromatin crescents, and pyknosis of the nuclei with some nuclear fragmentation. Meanwhile, three genes (API5, AIFM1, and NFκB1) showed changes of expression in the hippocampus of Ts65Dn mice compared with normal mice. Only NFκB1 expression was significantly increased, while the expressions of API5 and AIFM1 were notably decreased. The fold changes in the expression of API5, AIFM1, and NFκB1 were 11.55, 5.94, and 3.11, respectively. However, some well-known genes related to cell apoptosis, such as the caspase family, Bcl-2, Bad, Bid, Fas, and TNF, did not show changes in expression levels. The genes we found which were differentially expressed in the hippocampus of Ts65Dn mice may be closely related to cell apoptosis. PCR array technology can assist in the screening and identification of genes involved in apoptosis.

[Cu(H4C3N2S)Cl2](n), an unprecedented diazole-bridged one-dimensional copper halide: synthesis, structure, and magnetic properties.

The simultaneous desulfurization of 2-mercapto-5-methyl-1,3,4-thiadiazole with CuCl(2) x 2H(2)O via mutual diffusion in solvents results in the isolation of air-stable dark-green crystals of [Cu(H(4)C(3)N(2)S)Cl(2)](n) (approximately 65% yield). The structure is characterized by a unique one-dimensional copper chain bridged by diazine N-N single bonds rather than halogens, in sharp contrast with the halide bridging mode in conventional copper halide coordination polymers. Each Cu(II) ion shows a square planar coordination geometry featuring a strong Jahn-Teller distortion, as also supported by EPR data. The phase follows a Curie-Weiss paramagnetic behavior over 6-300 K. However, the intrachain antiferromagnetic interaction is evident (-2J = 21.1 cm(-1)). Such magnetic coupling is related to the interplay between the Cu(II)-d(x2-y2) and diazine N-N p-orbitals.