Neuroprotective effects of exosomes derived from bone marrow mesenchymal stem cells treated by Musk Ketone on ischemic stroke.

OBJECTIVES: Ischemic stroke (IS) is a leading cause of morbidity and mortality globally. This study aimed to investigate the role of exosomes (Exo) derived from bone marrow mesenchymal stem cells (BMSCs) treated with Musk Ketone (Mus treated-Exo) in the development of IS injury. METHODS: BMSCs were pretreated with 10 µM Mus for 36 h, and Exo derived from these Mus-treated BMSCs (Mus-treated Exo) were extracted. Rats with middle cerebral artery occlusion (MCAO) were administered either 2 mg/kg of control Exo (Ctrl-Exo), 2 mg/kg of Mus treated-Exo, or 10 µM Mus. Neurological deficit and cerebral infarction in the MCAO rats were assessed utilizing neurological scores and TTC staining. Neuronal apoptosis, activation of microglia/macrophages, and inflammation were evaluated through TUNEL staining, immunofluorescence staining, and western blot analysis, respectively. RESULTS: Our findings revealed that Mus-treated Exo possessed a more pronounced neuroprotective effect on MCAO rats when compared to Ctrl-Exo and Mus treatment alone. Specifically, Mus treated-Exo effectively ameliorated neurological function, reduced the volume of cerebral infarction, and diminished hemispheric swelling in MCAO rats. Moreover, it inhibited neuronal apoptosis and activation of microglia/macrophages, promoted the expression of the anti-apoptotic protein Bcl-2 while decreasing the expression of pro-apoptotic protein Bax, Cleaved-caspase 3, and pro-inflammatory factors IL-6 and COX-2. CONCLUSIONS: The findings imply that Mus treated-Exo could confer neuroprotection in rats affected by IS, potentially by attenuating apoptosis and neuroinflammation. The underlying mechanisms, however, warrant further investigation. Mus treated-Exo shows potential as a new therapeutic strategy for IS.

Therapeutic efficacy of tylvalosin combined with Poria cocos polysaccharides against porcine reproductive and respiratory syndrome.

Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically important infectious diseases of pigs worldwide. Vaccination and various management measures have been implemented to control PRRS. However, due to high genetic diversity and insufficient understanding of the pathogenesis and immunological mechanisms, PRRS is still a challenge to the pig industry. Therefore, it is important to develop novel strategies to combat PRRS virus (PRRSV) infection. In this study, our data show that tylvalosin, a third-generation animal-specific macrolide, could inhibit PRRSV replication in MARC-145 cells, and suppress the PRRSV-induced NF-κB activation and cytokines expression. The pig infection experiment further demonstrated that tylvalosin could significantly reduce the virus loads in serum and tissues, and alleviate lung lesions of pigs infected with highly pathogenic PRRSV strains. The fever and loss of daily gain (LoDG) of the pigs were decreased as well. Considering the feature of immune suppression of PRRSV, a combination of tylvalosin with the immunopotentiator Poria cocos polysaccharides (PCP) was developed. Pig experiment showed this combination had a better therapeutic efficacy against PRRSV infection than tylvalosin and PCP alone in attenuating lung lesions, alleviating fever, and suppressing cytokines production. This study suggests that tylvalosin has significant antiviral and anti-inflammatory effects against PRRSV infection, and the combination of tylvalosin and PCP provides a promising strategy for PRRS treatment.

Network-based drug repurposing for potential stroke therapy.

Stroke is the leading cause of death and disability worldwide, with a growing number of incidences in developing countries. However, there are currently few medical therapies for this disease. Emerged as an effective drug discovery strategy, drug repurposing which owns lower cost and shorter time, is able to identify new indications from existing drugs. In this study, we aimed at identifying potential drug candidates for stroke via computationally repurposing approved drugs from Drugbank database. We first developed a drug-target network of approved drugs, employed network-based approach to repurpose these drugs, and altogether identified 185 drug candidates for stroke. To validate the prediction accuracy of our network-based approach, we next systematically searched for previous literature, and found 68 out of 185 drug candidates (36.8 %) exerted therapeutic effects on stroke. We further selected several potential drug candidates with confirmed neuroprotective effects for testing their anti-stroke activity. Six drugs, including cinnarizine, orphenadrine, phenelzine, ketotifen, diclofenac and omeprazole, have exhibited good activity on oxygen-glucose deprivation/reoxygenation (OGD/R) induced BV2 cells. Finally, we showcased the anti-stroke mechanism of actions of cinnarizine and phenelzine via western blot and Olink inflammation panel. Experimental results revealed that they both played anti-stroke effects in the OGD/R induced BV2 cells via inhibiting the expressions of IL-6 and COX-2. In summary, this study provides efficient network-based methodologies for in silico identification of drug candidates toward stroke.

Apolipoprotein E mimetic peptide COG1410 alleviates blood­brain barrier injury in a rat model of ischemic stroke.

Blood­brain barrier (BBB) damage is one of the main causes of poor outcomes and increased mortality rates following cerebral ischemia­reperfusion injury. Apolipoprotein E (ApoE) and its mimetic peptide have been previously reported to exhibit potent neuroprotective properties in various central nervous system disease models. Therefore, the present study aimed to investigate the possible role of the ApoE mimetic peptide COG1410 in cerebral ischemia­reperfusion injury and its potential underlying mechanism. Male SD rats were subjected to 2 h middle cerebral artery occlusion followed by 22 h reperfusion. Evans blue leakage and IgG extravasation assays results revealed that COG1410 treatment significantly reduced BBB permeability. In addition, in situ zymography and western blotting were used to prove that COG1410 was able to downregulate the activities of MMPs and upregulate the expression of occludin in the ischemic brain tissue samples. Subsequently, COG1410 was found to significantly reverse microglia activation while also suppressing inflammatory cytokine production, according to immunofluorescence signal of Iba­1 and CD68 and protein expression of COX­2. Consequently, this neuroprotective mechanism mediated by COG1410 was further tested using the BV2 cell line in vitro, which was exposed to oxygen glucose deprivation followed by reoxygenation. The mechanism of COG1410 was found to be mediated, as least partly, through the activation of triggering receptor expressed on myeloid cells 2. In conclusion, the data suggest that COG1410 can alleviate BBB injury and neuroinflammation following ischemic stroke.

Systems Pharmacology-Based Strategy to Investigate the Mechanism of Ruangan Lidan Decoction for Treatment of Hepatocellular Carcinoma.

epatocellular carcinoma (HCC) is one of the leading contributors to cancer mortality worldwide. Currently, the prevention and treatment of HCC remains a major challenge. As a traditional Chinese medicine (TCM) formula, Ruangan Lidan decoction (RGLD) has been proved to own the effect of relieving HCC symptoms. However, due to its biological effects and complex compositions, its underlying mechanism of actions (MOAs) have not been fully clarified yet. In this study, we proposed a pharmacological framework to systematically explore the MOAs of RGLD against HCC. We firstly integrated the active ingredients and potential targets of RGLD. We next highlighted 25 key targets that played vital roles in both RGLD and HCC disease via a protein-protein interaction (PPI) network and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Furthermore, an ingredient-target network of RGLD consisting of 216 ingredients with 306 targets was constructed, and multilevel systems pharmacology analyses indicated that RGLD could act on multiple biological processes related to the pathogenesis of HCC, such as cellular response to hypoxia and cell proliferation. Additionally, integrated pathway analysis of RGLD uncovered that RGLD might treat HCC through regulating various pathways, including MAPK signaling pathway, PI3K/Akt signaling pathway, TNF signaling pathway, and ERBB signaling pathway. Survival analysis results showed that HCC patients with low expression of VEGFA, HIF1A, CASP8, and TOP2A were related with a higher survival rate than those with high expression, indicating the potential clinical significance for HCC. Finally, molecular docking results of core ingredients and targets further proved the feasibility of RGLD in the treatment of HCC. Overall, this study indicates that RGLD may treat HCC through multiple mechanisms, which also provides a potential paradigm to investigate the MOAs of TCM prescription.

The neuroprotective effect of YaoYi-moxibustion on ischemic stroke by attenuating NK-κB expression in rats.

Background: Traditional Chinese medicine (TCM) has become a crucial direction for ischemic stroke treatment. This study sought to explore the underlying roles of YaoYi-moxibustion (YY-moxi) in ischemic stroke. Methods: A total of 75 Sprague-Dawley rats were randomly divided into the following 5 groups: (I) the sham-operated group; (II) the middle cerebral artery occlusion model (MCAO) group; (III) the YY-moxi group; (IV) the antioxidant (N-acetylcysteine, NAC) group; and (V) the NAC + YY-moxi group. After the model had been established, the NAC group received intracerebroventricular injections of NAC, the YY-moxi group received YY-moxi, and the NAC + YY-moxi group received a combination of these 2 interventions. The neurological deficit score was confirmed, and the cerebral infarction was examined by triphenyl tetrazolium chloride (TTC) staining. In the ischemia site of stroke, terminal deoxynucleotidyl transferase-mediated Dutp nick end labeling staining was applied to examine the apoptotic cells. Additionally, the apoptosis-associated genes and protein expressions in the ischemic brains were investigated by the reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), immunohistochemistry, and western blot analysis. Results: YY-moxi alone and YY-moxi combined with NAC significantly reduced the neurological scores and cerebral infarction area of the MCAO rats. Additionally, YY-moxi alone and the combined application of YY-moxi and NAC improved the pathological status of ischemic brain tissues. Further, we found that YY-moxi alone and YY-moxi in combination with NAC could enhanced the antioxidation ability and reduced the inflammatory response of the MCAO model rats. We also proved that YY-moxi alone and YY-moxi combined with NAC significantly suppressed apoptosis-related proteins in the MCAO model rats. Conclusions: These findings indicate that YY-moxi exerts a protective effect on cerebral ischemic injury by reducing apoptosis. The study suggests that the mechanism may be related to its downregulating the expression of nuclear factor kappa B (NK-κB).

Low Expression of SLC7A11 Confers Drug Resistance and Worse Survival in Ovarian Cancer via Inhibition of Cell Autophagy as a Competing Endogenous RNA.

Drug resistance is the main cause of chemotherapy failure in ovarian cancer (OC), and identifying potential druggable targets of autophagy is a novel and promising approach to overcoming drug resistance. In this study, 131 genes associated with autophagy were identified from three autophagy-related databases, and of these, 14 were differentially expressed in 90 drug-resistant OC tissues versus 197 sensitive tissues according to the Cancer Genome Atlas ovarian cancer cohort. Among these 14 genes, SLC7A11 was significantly decreased in two paclitaxel-resistant OC cells (HeyA8-R and SKOV3-R) and in 90 drug-resistant tissues compared with their controls. In vitro overexpression of SLC7A11 significantly increased the sensitivity of HeyA8-R cells to paclitaxel, inhibited colony formation, induced apoptosis, and arrested cell cycle. Further, low SLC7A11 expression was correlated with poor overall survival (OS), progression-free survival (PFS), and post-progression survival (PPS) in 1815 OC patients. Mechanistically, SLC7A11 strongly regulated cell autophagy as a competing endogenous RNA (ceRNA) based on pan-cancer analyses of 32 tumor types. Specifically, as a ceRNA for autophagy genes STX17, RAB33B, and UVRAG, SLC7A11 was strongly and positively co-expressed with these three genes in 20, 12, and 12 different tumors, respectively, in 379 OC tissues and in 90 drug-resistant OC tissues, and the former two were significantly upregulated in SLC7A11-overexpressed HeyA8-R cells. Further, SLC7A11 induced the protein expression of other autophagy genes, such as LC3, Atg16L1, and Atg7, and the expression of the respective proteins was further increased when the cells were treated with paclitaxel. The results strongly suggest that SLC7A11 regulates autophagy via ceRNA interactions with the three abovementioned genes in pan-cancer and in drug-resistant OC. Moreover, low expression of STX17 and UVRAG also significantly predicted low OS, PFS, and PPS. The combination of SLC7A11 with STX17 was more predictive of OS and PFS than either individually, and the combination of SLC7A11 with UVRAG was highly predictive of OS and PPS. The above results indicated that decreased SLC7A11 resulted in drug resistance and effected low rates of survival in OC patients, probably via ceRNA interactions with autophagy genes, and thus the gene could serve as a therapeutic target and potential biomarker in OC.

NCALD affects drug resistance and prognosis by acting as a ceRNA of CX3CL1 in ovarian cancer.

Drug resistance, an impenetrable barrier in the treatment of ovarian cancer (OC), is often associated with poor outcomes. Hence, it is urgent to discover new factors controlling drug resistance and survival. The association between neurocalcin delta (NCALD) and cancer drug resistance is poorly understood. Here, we reveal that NCALD messenger RNA expression, probably regulated by DNA methylation and microRNAs, was significantly downregulated in at least three independent microarrays covering 633 ovarian carcinomas and 16 normal controls, which includes the Cancer Genome Atlas (TCGA) ovarian cohort. In the sub-groups of the TCGA cohort, NCALD was suppressed in 90 platinum-resistant tissues vs in 197 sensitive tissues. It is consistent with the quantitative reverse transcription polymerase chain reaction results revealing gene downregulation in carboplatin-resistant SKOV3 and HeyA8 OC cells as compared with that in controls. Low expression of NCALD predicted poor overall survival (OS) in sub-groups of 1656 patients, progression-free survival (PFS) in 1435 patients, and post-progression survival (PPS) in 782 patients according to Kaplan-Meier plotter covering 1815 OC patients. Comprehensive bioinformatic analyses strongly implicated NCALD in the regulation of drug resistance, probably via competing for endogenous RNA (ceRNA) interactions with CX3CL1 and tumor immune-microenvironment. NCALD acted as a ceRNA for CX3CL1 in 21 different cancers includes OC according to Starbase. These two genes negatively correlated with tumor purity and positively correlated with infiltration levels of neutrophils and dendritic cells in OC. The combined low expression of NCALD and CX3CL1 showed better prognosis potential for OS, PFS, and PPS in the 1815 OC patients than any of the individually tested genes. In summary, NCALD acts as a ceRNA for CX3CL1, and its downregulation may affect drug resistance and prognosis in OC. Thus, NCALD could be a new therapeutic target for anticancer therapy and a new biomarker for survival prediction in OC.

G1 and S phase arrest in Candida albicans induces filamentous growth via distinct mechanisms.

Candida albicans is an opportunistic fungal pathogen. In immunocompromised individuals, it can cause bloodstream infections with high mortality rates. The ability to switch between yeast and hyphal morphologies is a critical virulence factor of C. albicans. In response to diverse environmental cues, several signaling pathways are activated resulting in filamentous growth. Interestingly, cell cycle arrest can also trigger filamentous growth although the pathways involved are not well-understood. Here, we demonstrate that the cAMP-PKA pathway is involved in the filamentous growth caused by G1 arrest due to the depletion of the G1 cyclin Cln3 and S phase arrest due to hydroxyurea treatment. The downstream mechanisms involved in filamentation are different between the two cell cycle arrest phenomena. Cln3-depleted cells require HGC1 and UME6 for filamentous growth, but hydroxyurea-induced filamentation does not. Also, the hyphal repressor Nrg1 is not involved in the suppression of Cln3-depletion and hydroxyurea-induced filamentous growth. The findings highlight the complexity of the signaling networks that control filamentous growth in which different mechanisms downstream of the cAMP-PKA pathway are activated based on the nature of the inducing signals.

Cryptococcus strains with different pathogenic potentials have diverse protein secretomes.

Secreted proteins are the frontline between the host and pathogen. In mammalian hosts, secreted proteins enable invasive infection and can modulate the host immune response. Cryptococcosis, caused by pathogenic Cryptococcus species, begins when inhaled infectious propagules establish to produce pulmonary infection, which, if not resolved, can disseminate to the central nervous system to cause meningoencephalitis. Strains of Cryptococcus species differ in their capacity to cause disease, and the mechanisms underlying this are not well understood. To investigate the role of secreted proteins in disease, we determined the secretome for three genome strains of Cryptococcus species, including a hypovirulent and a hypervirulent strain of C. gattii and a virulent strain of C. neoformans. Sixty-seven unique proteins were identified, with different numbers and types of proteins secreted by each strain. The secretomes of the virulent strains were largely limited to proteolytic and hydrolytic enzymes, while the hypovirulent strain had a diverse secretome, including non-conventionally secreted canonical cytosolic and immunogenic proteins that have been implicated in virulence. The hypovirulent strain cannot establish pulmonary infection in a mouse model, but strains of this genotype have caused human meningitis. To directly test brain infection, we used intracranial inoculation and found that the hypovirulent strain was substantially more invasive than its hypervirulent counterpart. We suggest that immunogenic proteins secreted by this strain invoke a host response that limits pulmonary infection but that there can be invasive growth and damage if infection reaches the brain. Given their known role in virulence, it is possible that non-conventionally secreted proteins mediate this process.

The effect of standard heat and filtration processing procedures on antimicrobial activity and hydrogen peroxide levels in honey.

There is increasing interest in the antimicrobial properties of honey. In most honey types, antimicrobial activity is due to the generation of hydrogen peroxide (H(2)O(2)), but this can vary greatly among samples. Honey is a complex product and other components may modulate activity, which can be further affected by commercial processing procedures. In this study we examined honey derived from three native Australian floral sources that had previously been associated with H(2)O(2)-dependent activity. Antibacterial activity was seen in four red stringybark samples only, and ranged from 12 to 21.1% phenol equivalence against Staphylococcus aureus. Antifungal activity ranged from MIC values of 19-38.3% (w/v) against Candida albicans, and all samples were significantly more active than an osmotically equivalent sugar solution. All honey samples were provided unprocessed and following commercial processing. Processing was usually detrimental to antimicrobial activity, but occasionally the reverse was seen and activity increased. H(2)O(2) levels varied from 0 to 1017 µM, and although samples with no H(2)O(2) had little or no antimicrobial activity, some samples had relatively high H(2)O(2) levels yet no antimicrobial activity. In samples where H(2)O(2) was detected, the correlation with antibacterial activity was greater in the processed than in the unprocessed samples, suggesting other factors present in the honey influence this activity and are sensitive to heat treatment. Antifungal activity did not correlate with the level of H(2)O(2) in honey samples, and overall it appeared that H(2)O(2) alone was not sufficient to inhibit C. albicans. We conclude that floral source and H(2)O(2) levels are not reliable predictors of the antimicrobial activity of honey, which currently can only be assessed by standardized antimicrobial testing. Heat processing should be reduced where possible, and honey destined for medicinal use should be retested post-processing to ensure that activity levels have not changed.

[Transrectal ultrasound-guided puncture and anhydrous alcohol sclerotherapy for Müllerian duct cyst].

OBJECTIVE: To explore the effect of anhydrous alcohol sclerotherapy following transrectal ultrasound-guided puncture in the treatment of Mülllerian duct cyst. METHODS: Totally 44 patients with Müllerian duct cyst underwent transrectal ultrasound-guided puncture and sclerotherapy, 21 injected with anhydrous alcohol of half the volume of the aspirated cystic fluid followed by aspiration of all the fluid 5 minutes later (treatment group) , and the other 23 treated by cystic fluid aspiration only (control group). The clinical effects of the two methods were compared. RESULTS: The response rate and cure rate were 80.95 and 52.38% in the treatment group, as compared with 56.52 and 26.09% in the control (P < 0.001). No adverse events were observed in either of the two groups. CONCLUSION: Transrectal ultrasound-guided puncture and anhydrous alcohol sclerotherapy is a safe and effective approach to the treatment of Mülllerian duct cyst.

Pharmacokinetics and tissue residues of hydrochloric acid albendazole sulfoxide and its metabolites in crucian carp (Carassius auratus) after oral administration.

The pharmacokinetics and residues elimination of hydrochloric acid albendazole sulfoxide (ABZSO) and its metabolites were studied in healthy crucian carp (Carassius auratus, 250 ± 30 g) kept at water temperatures of 10 °C and 25 °C. The concentrations of ABZSO and its metabolites concentration in plasma and tissues were determined using high-performance liquid chromatography (HPLC) using an ultraviolet detector. The results revealed that the plasma concentration of ABZSO in plasma was significantly higher than that of albendazole sulfone (ABZSO(2)), whereas albendazole-2-aminosulfone (ABZ-SO(2)NH(2)) was not detected. The plasma concentrations of ABZSO and its main metabolite ABZSO(2) concentration-time data were fitted using a single-compartment model at 10 °C and 25 °C. The absorption half-life (t1/2ka) of ABZSO was 3.86 h at 10 °C and 1.29 h at 25 °C, whereas the elimination half-life (t1/2ke) was 16.34 h at 10 °C and 6.72 h at 25 °C; the maximum plasma concentration (C(max)) and the time-point of maximum plasma concentration (T(p)) were calculated as 3.20 µg mL(-1) and 10.58 h at 10 °C, 4.39 µg mL(-1) and 3.80 h at 25 °C. The distribution volume (V(d)/F) of ABZSO was estimated to be 1.99 L kg(-1) at 10 °C and 1.53 L kg(-1) at 25 °C; the total body clearance (CL(b)) of ABZSO were computed as 0.08 and 0.19 L/(h kg) at 10 and 25 °C, respectively; the areas under the concentration-time curve (AUC) was 118.22 µg mL(-1)h at 10 °C and 63.12 µg mL(-1)h at 25 °C. The [Formula: see text] of ABZSO(2) was found to be 6.39 °C at 10 °C and 3.73 h at 25 °C, whereas the [Formula: see text] was 12.86 h at 10 °C and 6.56 h at 25 °C; the C(max) and T(p) of ABZSO(2) was calculated as 0.78 µg mL(-1) and 12.82 h at 10 °C, 1.03 µg mL(-1) and 7.04 h at 25 °C, respectively; the V(d)/F of ABZSO(2) were estimated to be 6.43 L kg(-1) at 10 °C and 4.61 Lkg(-1) at 25 °C; the CL(b) of ABZSO(2) were computed as 0.34 and 0.49 L/(h kg) at 10 °C and 25 °C, respectively; the AUC of ABZSO(2) were 28.86 µg mL(-1)h at 10 °C and 20.52 µg mL(-1)h at 25 °C. It was demonstrated that ABZSO(2) was the main metabolite of ABZSO. The concentrations of ABZSO and its main metabolite (ABZSO(2)) were detected in muscle, skin, liver and kidney, whereas ABZ-SO(2)NH(2) was only detected in liver and kidney. The ABZSO and it metabolite (ABZSO(2)) could still be detected at 4 d time-point after administration at both temperatures in all tissues. The results revealed that the depletion of ABZSO and its metabolite (ABZSO(2)) in crucian carp was slower with a long half-life time, especially at lower water temperature.

Preparation and evaluation of danofloxacin mesylate microspheres and its pharmacokinetics in pigs.

Danofloxacin mesylate gelatin microspheres (DFM-GMS) were prepared by an emulsion chemical crosslinking technique. Distribution of particle size, morphologic characteristics, drug content, and drug stability were evaluated. In-vitro study showed that the release of danofloxacin mesylate (DFM) from microspheres was much slower than from the raw material (DFM) in the release medium. Pharmacokinetic characteristics were evaluated following intramuscular injection of DFM-GMS or DFM in pigs at dosage of 2.5 mg/kg body weight. Elimination half-life (t(1/2ß)) of the drug was 24.32 h for DFM-GMS, and 6.61 h for DFM (P < 0.01). Overall, DFM-GMS could be applied as a long-acting and lung targeting dosage form of DFM for clinical application.