Anti-fungal efficacy of combination of amphotericin B with colistin and gentamicin in McCarey-Kaufman corneal preservation medium

Purpose: To curtail the potential of donor corneal tissue disseminating fungi to the recipient抯 eye, we evaluated the addition of amphotericin B to McCarey?Kaufman (M?K)梒orneal storage medium supplemented with colistin. Methods: Amphotericin B was examined for its ability to inhibit the growth of Candida albicans and Aspergillus flavus using a microbroth dilution test and checkerboard assay in combination with only gentamicin and a combination of colistin, gentamicin, and amphotericin B. The safety on epithelium and endothelium was evaluated by 3?(4,5?dimethylthiazol?2?yl)?2, 5?diphenyltetrazolium bromide (MTT) assay. Results: The minimal inhibitory concentration of gentamicin was found to be >256 ?g/ml against both C. albicans and A. flavus, whereas that of amphotericin B was found to be in a range of 0.25�5 and 1�?g/ml for C. albicans and A. flavus, respectively. According to the checkerboard assay, 80% (4/5) of C. albicans isolates and 100% (5/5) of A. flavus isolates responded synergistically to the combination of amphotericin B and gentamicin, but only 20% (1/5) of C. albicans isolates showed an additive effect. None of the tested isolates displayed antagonism. The combined effect of the three drugs also did not display any antagonistic effect. Additionally, the MTT assay reveals no toxic effect of the antimicrobials used on corneal epithelial and endothelial cells. Conclusion: In vitro experiments demonstrate that amphotericin B is not toxic to either epithelium or endothelium and is a promising additive to the M?K medium supplemented with colistin.

The role of NMDA glutamate receptors in lung injury caused by chronic long-term intermittent hypobaric hypoxia

Chronic intermittent hypoxia (CIH), a component of sleep apnea-hypopnea syndrome, is suggested to cause damage to lung tissue, and the role of glutamate is not well studied. We used a chronic long-term intermittent hypobaric hypoxia (CLTIHH) model of rats to find out if such procedure causes lung injury and the potential effect of N-methyl-D-aspartate receptors (NMDARs) by using receptor antagonist MK-801 (dizocilpine). Thirty-two rats were placed into four groups; a control and three CLTIHH groups where rats were placed into a low-pressure chamber set to 430 mmHg for 5 h/day, 5 days/week, for 5 weeks. Only one group received MK-801 (0.3 mg/kg, ip) daily. We evaluated tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-10, and nuclear factor (NF)-kB for the inflammatory process, superoxide dismutase (SOD), malondialdehyde (MDA), catalase (CAT), glutathione peroxidase (GPX), total antioxidant status (TAS), and total oxidant status (TOS) for oxidative stress, and caspase-9 levels. Blood plasma, bronchoalveolar fluid (BALF), and lung tissue extracts were evaluated. Both oxidant and inflammatory parameters were significantly increased in all the mediums of the CLTIHH groups except the group that received MK-801. Significant evidence was collected on MK-801 alleviating the effect of CLTIHH. Histological evaluations revealed lung damage and fibrotic changes in the CLTIHH groups. It was first shown that the CLTIHH procedure caused chronic lung injury, and that inflammation and oxidant stress were influential in the formation of lung injury. Secondly, NMDAR antagonist MK-801 effectively inhibited the development of lung injury and fibrosis.

Research progress of methodological techniques of generation of platelets from induced pluripotent stem cells in vitro / 中国输血杂志

Platelets play a role in hemostasis in vivo, and platelet transfusion is the main means to treat bleeding diseases caused by thrombocytopenia or platelet dysfunction. However, platelets are in short supply due to the increasing demand for platelet products in clinical, the limited number of blood donors and the disadvantages of platelet products such as short shelf life and bacteria contamination. Currently, induced pluripotent stem cells are considered an ideal source for producing platelets in vitro. They have the potential for self-renewal and differentiation into any cell type, and can be obtained and manipulated easily. Given the recent advances in megakaryocytic series, bioreactors, feeder-free cell production and large-scale propagation research, platelet preparations derived from induced pluripotent stem cells have gradually shown great potential for clinical applications. Considering the minimal risk of alloimmunization and tumorigenesis with these blood products, they are promising to become the standard source of future blood transfusions. This paper reviews the research progress of the methodological techniques of in vitro generation of platelets from induced pluripotent stem cells.

An experimental study to evaluate the effect of polymixin E (Colistin) alone or in combination with gentamicin in McCarey-Kaufman corneal preservation medium on various drug resistant bacterial and fungal isolates

Purpose: To assess the efficacy of the addition of polymyxin E (colistin) in the McCarey-Kaufman (MK) corneal storage solution against multi-drug resistant strains of Enterobacteriaceae, Staphylococcus aureus, and Candida spp. Methods: A standard micro broth dilution test and a checkerboard assay were performed for five multi-drug resistant (MDR) clinical strains of P. aeruginosa and five clinical strains of methicillin-resistant S. aureus (MRSA) and C. albicans against colistin and gentamicin alone and in combination. The minimum inhibitory concentration (MIC) and the fractional inhibitory concentration index (FICI) were calculated to assess the efficacy of each combination. Results: The MIC of colistin was in the range of 1–2 ?g/mL for P. aeruginosa, whereas it was 256–1024 ?g/mL against S. aureus. In comparison, the MIC of gentamicin was found to be 0.5–512 ?g/mL and 0.5–8 ?g/mL against P. aeruginosa and S. aureus, respectively. All five isolates of C. albicans did not exhibit any susceptibility to either colistin or gentamicin even at a concentration of ? 512 ?g/ mL each. The checkerboard assay was performed to evaluate the nature of the interaction of the combination of colistin and gentamicin. Based on the FICI, it was observed that the colistin and gentamicin combination has a maximum synergistic effect (FIC <0.5) in 80% (4/5) for S. aureus isolates, whereas the maximum additive effect (FIC >0.5–4) was 100% (5/5) for P. aeruginosa and the minimum additive effect was 20% (1/5) for S. aureus isolates. Antagonism (FIC ? 4) was not observed in any combination between the strains used in the study. Both colistin and gentamicin alone or in combination were, however, ineffective against Candida spp. Conclusion: The addition of colistin has an inhibitory effect on bacterial contamination that could be possibly caused by MDR strains and could potentially be considered as an additional additive in corneal storage media.

Efficacy of voriconazole and amphotericin B in corneal preservative media

Purpose: To evaluate the efficacy of voriconazole and amphotericin B in McCarey–Kaufman (MK) media. Methods: MK media vials were supplemented with either voriconazole at 1, 2, 20, 50, 100 ?g/mL or amphotericin B at 0.5, 1, 2, 10, 20 ?g/mL. The standard inoculum of the American Type Culture Collection (ATCC) strain of Candida albicans, Aspergillus flavus, and Fusarium keratinoplasticum was added to the set of vials. The efficacy outcomes were calculated as ‘viable fungal colony counts’ determined from the samples taken on Days 0 and 4. MK media containing fungal inoculum but without antifungal supplements were used as control. Results: In the voriconazole arm, on Day 4, a reduction in the colony count was observed for Candida albicans (1 ?g/mL, 36%; 100 ?g/mL, 100%), Aspergillus flavus (1 ?g/mL, 53.8%; 100 ?g/mL, 80.4%), and Fusarium keratinoplasticum (1 ?g/mL, 39.0%; 100 ?g/mL, 72.2%). Similarly, in the amphotericin B arm, on Day 4, a reduction in the colony count was observed for Candida albicans (0.5 ?g/mL; 99.9%; 20 ?g/mL, 100%), Aspergillus flavus (0.5 ?g/mL, 65.2%; 20 ?g/mL, 84.8%), and Fusarium keratinoplasticum (0.5 ?g/mL, 90.1%; 20 ?g/mL, 100%). Conclusion: Compared to voriconazole, the addition of amphotericin B significantly reduces fungal contamination in MK media.

Mogroside V inhibits H202-induced oxidative damage in MIN6 cells by PI3K/Akt pathway / 中国药理学通报

Aim To investigate the protective effect of mogroside V on hydrogen peroxide ( H,02 )-induced oxidative stress response in mouse islet (3 cells MIN6 and the relation of its mechanism to PI3K/Akt signa¬ling pathway.Methods MIN6 cells were treated with 500 (jimol • L_1 H,(), after mogroside V,and cell via¬bility was detected by MTT.The release of reactive ox¬ygen species ( ROS) and apoptotic percentage of MIN6 cells were determined by flow cytometry.The expres¬sions of apoptosis-related factor Bel-2 , proliferation-re¬lated factor PCNA, protein Akt and p-Akt were deter¬mined by Western blot.Results H,02 restrained the proliferation of MIN6 cells obviously, induced ROS pro¬duction and apoptosis, and reduced the expression of Bel-2 and PCN A.The expressions of protein Akt and p-Akt decreased.After treatment of mogroside V , the release of ROS decreased, and the apoptosis of MIN6 cells was inhibited.The expression levels of apoptosis- related protein Bcl-2 and proliferation-related protein PCN A were reversed.The expressions of protein Akt and P-Akt increased.The viability of MIN6 cells in¬duced by H,0, increased.In addition, mogroside V partly reversed the apoptosis induction and ROS pro¬duction of Akt inhibitor MK2206 (5 jjimol • L"1 ) on MIN6 cells.Conclusions Mogroside V has protec¬tive effect on H202-induced oxidative damage in MIN6 cells and its mechanism is related to PI3K/Akt signa¬ling pathway.

Spatiotemporal distribution of oncomelania hupensis snail habitats in the Middle and Lower Reaches of the Yangtze River / 公共卫生与预防医学

Schistosomiasis is one of the major communicable diseases, which seriously impacts human health, Oncomelania hupensis snail is unique intermediate host of Schistosoma japonicum as well as posts significant influence on schistosomiasis transmission. The long-term serial data of oncomelania snail area in the Middle and Lower Reaches of the Yangtze River in the Schistosomiasis situation in People's Republic of China ,were collected from 1999 to 2018. The spatial and temporal variation characteristics of oncomelania hupensis area in five provinces were analyzed by Manner-Kendall test. In the spatial change, the area composition ratio of lake-and-marshland regions in Hunan increased, while that in Hubei and Jiangxi showed a slight decline. The lake-and-marshland snails in Anhui showed a significant downward trend, while Jiangsu showed no stable trend. In the spatial change, the area of lake-and-marshland snails in Jiangsu, Anhui, and Hubei showed a downward trend, among which the downward trend in Hubei was not obvious, while the upward trend in Jiangxi and Hunan had an upward trend but the trend was not obvious. the year of the sudden change in the area of the lake and marshes in in Jiangsu and Hubei were 2010 and 2017 ; There was no significant mutation in Anhui Province; the mutation point of the lake-and-marshland snails in Hunan was between 2001 and 2002; the mutation year of the lake-and-marshland snails in Jiangxi was 2002.This paper studies the temporal and spatial changes of the snail breeding area in the middle and lower Yangtze River, and the results can provide a basis for the elimination of snails in different types of distribution areas.

Xinfeng Capsules promotes apoptosis of synovial fibroblasts and attenuates inflammation in rheumatoid arthritis by regulating lncRNA MAPKAPK5-AS1 / 中国中药杂志

To explore the regulatory effects of Xinfeng Capsules(XFC) on the apoptosis of synovial fibroblasts(FLS) and inflammation in rheumatoid arthritis(RA) via lncRNA MAPKAPK5-AS1(MK5-AS1). Thirty healthy people and 30 patients with RA due to spleen deficiency and dampness exuberance were collected for extracting the peripheral blood mononuclear cells(PBMCs) before and after XFC treatment, which were used to observe the correlation between MK5-AS1 and clinical indicators as well as MK5-AS1 expression before and after XFC treatment. Following the establishment of RA-FLS cell line and the preparation of XFC-containing serum, MK5-AS1-overexpression plasmid was constructed and transfected into RA-FLS for investigating the efficacy of XFC-containing serum in regulating inflammation and apoptosis of RA-FLS via MK5-AS1. The expression of MK5-AS1 in PBMCs of patients with RA due to spleen deficiency and dampness exuberance was decreased(P<0.001). The ROC curve analysis revealed the AUC of 83.9%. Correlation analysis showed that MK5-AS1 was negatively correlated with ESR, CRP, RF, CCP, and spleen deficiency and dampness exuberance syndrome score. The expression of MK5-AS1 increased significantly after XFC treatment(P<0.001). As demonstrated by association analysis, XFC decreased MK5-AS1, ESR, CRP, RF, and spleen deficiency and dampness exuberance syndrome score, with the degree of support all greater than 83%, confidence greater than 80%, and lift greater than 1. The results of RT-qPCR showed that the MK5-AS1 RNA expression significantly decreased after TNF-α stimulation(P<0.01), which, however, increased significantly after the intervention with XFC-containing serum(P<0.05). Such expression rose again after the transfection of pcDNA3.1-MK5-AS1(P<0.01). ELISA results showed that TNF-α stimulation elevated the expression of pro-inflammatory factor IL-17 but lowered the expression of anti-inflammatory factor IL-4(P<0.01). After intervention with XFC-containing serum, the expression of IL-17 decreased while that of IL-4 increased(P<0.01). The transfection of pcDNA3.1-MK5-AS1 contributed to the reduction in IL-17 expression but the elevation in IL-4 expression(P<0.01). The immunofluorescence(IF) findings demonstrated that the expression of pro-apoptotic protein Bax was down-regulated, whereas that of the anti-apoptotic protein Bcl-2 was up-regulated after TNF-α stimulation(P<0.01). After the intervention with XFC-containing serum, the Bax expression was increased, while Bcl-2 expression was decreased(P<0.01), which were remarkably collaborated by the transfection of pcDNA3.1-MK5-AS1(P<0.05). The expression of MK5-AS1 is significantly decreased in both RA-PBMCs and RA-FLS, implying that XFC inhibits inflammatory reaction and promotes the apoptosis in RA by regulating the expression of MK5-AS1.

Experimental study of diffusion kurtosis imaging in predicting radiotherapy sensitivity of esophageal carcinoma / 中华放射医学与防护杂志

Objective To explore the value of diffusion kurtosis imaging (DKI) in predicting radiotherapy sensitivity of esophageal cancer from the animal model level.Methods BALB/c nude mice were subcutaneously injected with Eca-109 cell lines to form xenograft tumors.The tumors received a single dose of 15 Gy (6 MV X-rays) in the experimental group or had no any treatment as control.The volume of transplanted tumor,the change of ADC,MK and MD values,and the tumor cell density and necrosis ratio of these two groups were observed at the corresponding time points.Results The growth of xenograft volume in the experimental group was suppressed and it was significantly smaller than that in the control group (t=3.206-6.149,P＜0.05) at the 7th day after radiotherapy.From the 3rd day after radiotherapy,the ADC and MD values of the experimental group were significantly higher than those of the control group,and the MK values was lower than those in the control group (tADC =-11.018--2.049,tMD =-6.609--2.052,tMK =2.492-9.323,P＜0.05).Meanwhile,the tumor cell density of the control group was higher than that of the experimental group,and the proportion of necrosis in the experimental group was higher than that in the control group (tdensity =-8.387--2.239,t is =2.980-17.430,P＜0.05).Conclusions A single large dose radiation could inhibit the growth of xenograft.ADC,MK,MD values changed at the early stage prior to morphological changes of tumor in consistent with the change of cell density and necrosis ratio.DKI has the potential value in predicting radiotherapy sensitivity of esophageal carcinoma.

NMDA Receptor Antagonist MK801 Protects Against 1-Bromopropane-Induced Cognitive Dysfunction / 神经科学通报·英文版

Occupational exposure to 1-bromopropane (1-BP) induces learning and memory deficits. However, no therapeutic strategies are currently available. Accumulating evidence has suggested that N-methyl-D-aspartate receptors (NMDARs) and neuroinflammation are involved in the cognitive impairments in neurodegenerative diseases. In this study we aimed to investigate whether the noncompetitive NMDAR antagonist MK801 protects against 1-BP-induced cognitive dysfunction. Male Wistar rats were administered with MK801 (0.1 mg/kg) prior to 1-BP intoxication (800 mg/kg). Their cognitive performance was evaluated by the Morris water maze test. The brains of rats were dissected for biochemical, neuropathological, and immunological analyses. We found that the spatial learning and memory were significantly impaired in the 1-BP group, and this was associated with neurodegeneration in both the hippocampus (especially CA1 and CA3) and cortex. Besides, the protein levels of phosphorylated NMDARs were increased after 1-BP exposure. MK801 ameliorated the 1-BP-induced cognitive impairments and degeneration of neurons in the hippocampus and cortex. Mechanistically, MK801 abrogated the 1-BP-induced disruption of excitatory and inhibitory amino-acid balance and NMDAR abnormalities. Subsequently, MK801 inhibited the microglial activation and release of pro-inflammatory cytokines in 1-BP-treated rats. Our findings, for the first time, revealed that MK801 protected against 1-BP-induced cognitive dysfunction by ameliorating NMDAR function and blocking microglial activation, which might provide a potential target for the treatment of 1-BP poisoning.

Recapitulation of Neuropsychiatric Behavioral Features in Mice Using Acute Low-dose MK-801 Administration

Despite some innate limitations, animal models are a potent investigative tool when used to model specific symptoms of a disorder. For example, MK-801, an N-methyl-D-aspartate receptor antagonist, is used as a pharmacological tool to induce symptoms found in some neuropsychiatric disorders. However, a close examination of literature suggests that the application window of MK-801 doses is relatively narrow between individual behavioral paradigms, necessitating careful characterization of the evoked behavioral aberrations and the doses used to induce them. Moreover, variation in behaviors depending on the animal strain, gender of the subject, and the timing of administration is observed, making it difficult to compare the behavioral characteristics reported in different studies. We aim to characterize the behavioral aberrations induced by different doses of MK-801 in CD-1 mice and create a ready reference for future studies. We used CD-1 mice to recapitulate behavioral impairments resulting from acute administration of MK-801. In 0.1 mg kg⁻¹, we observed diminished spontaneous alteration during the Y-maze test, while 0.12 mg kg⁻¹ resulted in hyperlocomotion and social deficit. Mice treated with 0.2 and 0.3 mg kg⁻¹ of MK-801 demonstrated a decreased self-grooming. Finally, all doses significantly impaired cliff avoidance behaviors suggesting increased impulsivity. These results affirm that MK-801 can effectively model various symptoms of different neuropsychiatric disorders in a dose-dependent manner. The observed sensitivity against spatial-memory impairment and impulsive behaviors at low concentration of MK-801 suggest that MK801 may modulate cognitive function and impulsivity in even lower concentration before it can modulate other behavioral domains.

α-humulene improves schizophrenia via NRG1/ErbB4 signaling pathway / 中草药

Objective: To study the treating mechanism of α-humulene on the schizophrenic mice. Methods: The schizophrenic models were established by dizocilpine maleate (MK801), then different concentrations of α-humulene were used to treat the mice by intragastric administration. Open-field experiment and PPI test were carried out to evaluate the spontaneous activity and sensorimotor gating function of mice. Moreover, the frontal cortex MDA, NO levels and hippocampal NRG1, ErbB4 protein expression was detected. Results: The spontaneous activity, sensorimotor gating function, MDA, NO, NRG1 and ErbB4 levels were significantly changed in model mice when compared with normal mice (P < 0.01). Compared with model group, different concentrations of α-humulene notably inhibited spontaneous activity, improved PPI value, increased NO and MDA content, down-regulated ErbB4 and NRG1 protein expression (P < 0.05, P < 0.01). Conclusion: The schizophrenia abnormal behavior of mice was improved by α-humulene via down-regulating NRG1/ErbB4 signaling pathway, so as to achieve the purpose of treating schizophrenia.

Effect of dizocilpine (MK-801) on the working memory of rats on a three-panel runway apparatus

Background: Understanding the processes underlying cognitive functions is a prerequisite to develop strategies for the treatment of cognitive deficits. There is a great need for valid animal models for investigating the cognitive enhancing effects of potential therapeutics. Many studies have investigated animal models of cognitive deficits by using animals treated with compounds that compromise cognitive abilities. Glutamate, an excitatory neurotransmitter and abundantly distributed in the central nervous system is involved in memory processes through N-methyl-d-aspartate (NMDA) receptors. The behavioural consequences of blocking the NMDA receptor provide the rationale for cognitive impairment as an animal model for the cognitive deficits associated with dementia. Authors investigated the effect of dizocilpine (MK-801), an NMDA-receptor antagonist (non-competitive) on the working memory in rats using the three-panel runway apparatus.Methods: Total 24 trained male albino rats were randomly divided into 4 groups of 6 animals each. Varying doses of MK-801 were administered to the animals. Working memory errors and latency periods were evaluated on the three panel Runway apparatus.Results: Treatment with MK-801 at the dose of 0.03mg/ kg did not result in any significant change in working memory errors or latency period in comparison to saline control. MK-801 treatment at dose of 0.1mg/kg and 0.3mg/kg resulted in a significant increase in the number of working memory errors and latency period as compared to control.Conclusions: Authors conclude that MK-801 treatment in the dose of 0.1mg/ kg and 0.3mg/kg resulted in working memory deficits on the three-panel runway apparatus. Rats with cognitive deficits induced by the prototypical N-methyl-d-aspartate (NMDA) receptor antagonist MK-801 may provide a relevant animal model of dementia based on the mechanistic approach of blocking NMDA/glutamatergic signalling.

Effects of neonatal MK-801 exposure on recognition memory and excitatory-inhibitory balance of hippocampus in adult female rats / 中华行为医学与脑科学杂志

Objective To investigate the long-term effects of repeated neonatal administration of dizocipline maleate (MK-801),the glutamate N-methyl-D-aspartate (NMDA) receptor antagonist,on recognition memory and hippocampal excitatory-inhibitory balance at the synaptic level in adult female rats.Methods Neonatal female Sprague-Dawley (SD) rats were randomly divided into model group and control group.Rats were administrated subcutaneously with MK-801 or normal saline from postnatal day (PND) 5 to PND14 (0.25 mg/kg,twice daily).(1) Object-in-context recognition test was performed on PND73-75.(2)The expression levels of vesicular glutamate transporter 1 (VGLUT1) and vesicular GABA transporter (VGAT) in hippocampus were detected by immunohistochemical staining.Results (1) The preference index of model group for new objects was significantly lower than that of the control group (t =-2.762,P=0.012).(2) There was no significant difference in the expression of VGLUT1 in hippocampus of MK-801 mode group(P＞0.05).Compared with control group(48.19±2.10),the VGAT level of model group in CA1 (39.60±2.19) was lower.Compared with control group (CA1:(0.99±0.05),CA3:(1.28±0.02),the ratio of VGLUT1/VGAT was significantly upregulated in CA1(1.16±0.05) and CA3(1.44±0.03) (P＜0.05).Conclusion Early NMDA receptor inhibition produces long-term deleterious effects on associative recognition memory and excitatory-inhibitory balance of hippocampus in female rats.These biochemical abnormalities may contribute to cognitive impairments observed in this study.

Role of Akt during Islet-1 inducing C3H10T1/2 cells to differentiate specifically into cardiomyocytes / 基础医学与临床

Objective To study the effect and mechanism of Akt during Islet -1 inducing the mesenchymal stem cells of C3H10T 1/2 cells to differentiate specifically into cardiomyocytes.Methods Developing a model of Islet-1 over-expression cells, Lentivirus infection efficiency of cells was detected by flow cytometry detection tech-nology.Cell proliferation was tested by CCK-8.The protein expression of Islet-1, cTnT and p-Akt/T-Akt was measured by Western blot.The mRNA expression of GATA4,Nkx2.5 and Mef2c were tested by real-time PCR. Results With the increasing of MK-2206 concentration, the inhibition rate of cell proliferation increased ( P<0.05) ,and the best inhibition concentration of Akt was 8nmol /L; With prolongation of Islet-1 inducing time,the protein expression of p-Akt/TA-kt reduced ( P<0.05 );The gene expression of myocardial specific transcription factor GATA4, Nkx2.5 andMe f2c increased( P<0.05); treated with MK-2206, the geneexpres-sion of GATA4, Nkx2.5 and Mef2cincreased significantly at the first week and then reduced ( P<0.05) . Conclu isons Akt plays different effects in different differentiation stages during the Islet-1 inducing the cells into cardiacs-pecific differentiation process .

Radiation dose effect on non-small cell lung cancer based on driver genotyping: An exploratory study / 肿瘤

Objective: To explore the importance of distinguishing α/β values for different lung cancer cells based on driver genes and its significance in the precise radiotherapy. At the same time, to study the influence of serine/threonine kinase inhibitor MK1775 on the α/β values for different kinds of cells, and to investigate the radiation sensitization or synergistic effect of MK1775. Methods: Lung cancer PC9 cells with epidermal growth factor receptor (EGFR) gene mutant, A549 cells with V-Ki-ras2-Kirsten rat sarcoma viral oncogene homolog (K-RAS) gene mutant, and H1299 cells with p53 gene mutant were respectively cultured and assigned into three groups: the control group, radiation alone group, and MK1775 inhibitor combined with radiation group. 6 MV X-ray with a dose rate of 3 Gy/min was employed in all the radiation groups, in which the administered dosages were 0, 0.25, 0.5, 1, 2, 3, 4, 5 and 6 Gy, respectively. Finally, the dose-effect relationship was analyzed by employing clone counting method, and the radiosensitization and synergistic effects of MK1775 on the three kinds of gene mutant cells were analyzed. Results: The different gene mutation types corresponded to different hyper-radiosensitivity doses. The numbers of PC9, A549 and H1299 cell clones were decreased to 56.4%, 54.5% and 73.2% respectively after adding MK1775 inhibitor, which indicated that the synergy effect exists indeed between MK1775 and radiation. The α/β values were negative in PC9 and H1299 cells (except of A549 cells) before MK1775 treatment, and all the α/β values in PC9, H1299 and A549 cells in MK1775 combined with radiation group were positive. So it was indicated that the radiosensitizing effect of MK1775 inhibitor only worked when α/β value was negative. Additionally, there was significant difference between the radiation alone group and MK1775 combined with radiation group (P < 0.05). Conclusion: The different driver genes correspond to different α and β values, thus correspond to different equivalent biological doses. Simultaneously, MK1775 maybe have the different radiosensitization or radiosynergistic effect for lung cancer cells with different driver genotypes.

Isolation, Characterization, Antioxidant, Antimicrobial and Cytotoxic Effect of Marine Actinomycete, Streptomyces Carpaticus MK-01, against Fish Pathogens

ABSTRACT Present study aim to evaluate the antimicrobial, antioxidant and cytotoxic potential of crude extract of Marine Streptomyces carpaticus MK-01 isolated from seawater collected from Daejeong-cost of Jeju Island. About 24 actinomycetes strains were isolated and subjected to morphological and molecular analysis that confirmed the isolate as S. carpaticus MK-01. Crude ethyl acetate extract of MK-01 strain showed extensive antibacterial activity against Gram-positive fish pathogenic bacteria namely Streptococcus iniae and S. parauberis with a maximum zone of inhibition (0.92±0.03mm) was recorded against S. parauberis at the minimum extract concentration (3.12µg/ml). The MK-01 ethyl acetate extract shows dose dependant significant increase in antioxidant activity. The 50% cytotoxic concentration (CC50) of MK-01 ethyl acetate extract was attained at 53.71 μg/ml and the effective concentration 50 (EC50) against virus-infected Epithelioma papulosum cyprini cell lines was 8.72 μg/ml of S. carpaticus MK-01 crude ethyl acetate extract.

Fraccionamiento guiado por bioensayo e identificación de blancos moleculares de potencial acción antipsicótica de cuatro plantas medicinales peruanas / Fractionation guided by bioassay and identification of molecular targets of potential antipsychotic action of four Peruvian medicinal plants

El principal síntoma de la esquizofrenia es la psicosis, caracterizada principalmente por la aparición de alucinaciones y delusiones. En esta investigación, se llevó a cabo el fraccionamiento biodirigido de extractos etanólicos (EE) de 4 plantas (6078, 6518, 7158 y 6521) usadas en la medicina tradicional Peruana para el tratamiento de psicosis. Los extractos y fracciones fueron probados en el modelo animal murino de hiperactividad inducida por Dizocilpina o MK-801(antagonista del receptor de glutamato tipo N-metil D-aspartato). Se utilizó la prueba de campo abierto (OFT) para medir la hiperactividad y la prueba de inhibición del sobresalto por prepulso (PPI) para medir la respuesta de sobresalto. En la prueba de campo abierto, la fracción apolar A del EE 7158 (p<0.05), la fracción D del EE 6078 (p <0.001) y la fracción A (p<0.05) y D (p<0.001) del EE 6518 (p<0.001) suprimieron los efectos del MK-801. En la prueba de PPI fueron activas las fracciones A, C y D (7158); A, B y C (6078); y todas las fracciones del extracto 6518. Adicionalmente, los ensayos de unión a radioligando y ensayos funcionales indican que los cuatro EEs tienen un efecto sobre receptores involucrados en esquizofrenia.

Effect of Artesunate on Akt/GSK-3β/β-catenin Signal Pathway in Human Hepatic Stellate Cells / 中国药师

Objective: To explore the effect of artesunate (Art) on Akt/GSK-3β/β-catenin signal pathway.Methods: Art at different concentrations (0, 12.5, 25, 50 μg·ml-1) was used to treat human hepatic stellate cells (LX-2), and CCK-8 assay was used to detect the cell proliferation to determine the optimal concentration.Art inhibitor (MK-2206) at different concentrations (0~8 μmol·L-1) was given to LX-2 cells, and a Western Blot method was applied to determine the optimal inhibition concentration.Art, MK-2206 and MK-2206+Art were respectively given to LX-2 cells, and a Western Blot method was used to detect the levels of Akt, p-Akt, GSK-3β, p-GSK-3β and β-catenin proteins.Results: CCK-8 assay was used to detect the cell survival rate, and the survival rate was 80% after the 24-hour treatment with 25 μg·ml-1 Art.The results of Western Blot showed that MK-2206 at 6 μmol·L-1 could effectively inhibit the expression of p-Akt.Compared with those of the control group, the levels of Akt, p-Akt, p-GSK-3β and β-catenin protein were significantly different (P＜0.05) in Art (25 μg·ml-1) group, MK-2206 (6 μmol·L-1) group and MK-2206 (6 μmol·L-1) + Art (25 μg·ml-1) group.The expression of GSK-3β and Akt in MK-2206+Art group had no significant difference when compared with that in Art group and MK-2206 group (P＞0.05), while the levels of p-Akt, p-GSK-3β and β-catenin were significantly reduced (P＜0.01).Conclusion: Art exhibits the influence on the relative factors in Wnt/β-catenin signal pathway by Akt/β-catenin, subsequently inhibits the cell proliferation and alleviates the liver fibrosis process.

Combination of everolimus and MK-2206 for synergistic inhibition of hepatocarcinoma cell proliferation / 中国药理学与毒理学杂志

OBJECTIVE To examine the synergistic inhibiory effect of combination of mammalian target of sirolimus (Rapamycin) (mTOR) inhibitor everolimus and AKT inhibitor MK-2206 on hepatocar-cinoma cell proliferation. METHODS HepG2 and BEL-7402 cells were treated with sirolimus and evero-limus alone for 0, 1, 3, 6, 12 and 24 h or in combination with insulin-like growth factor 1 receptor (IGF-1R) inhibitor NVP-AEW541 or AKT inhibitor MK2206 for 24 h. p70S6K and AKT kinase activityies were detected by Western blotting. Plate clone formation assay and CCK8 assay were used to detect the growth and proliferation of hepatocarcinoma cells treated with everolimus and MK2206 alone or in combi-nation. RESULTS Sirolimus and everolimus inhibited p70S6K activity while causing feedback activa-tion of AKT kinase activity at different time points (P<0.01). NVP-AEW541 and MK-2206 could inhibit AKT kinase feedback activation by everolimus (P<0.05). Colony formation of hepatocarcinoma cells treated with everolimus and MK-2206 in combination was significantly inhibited compared with everolimus or MK-2206 alone (P<0.01). Everolimus and MK-2206 in combination inhibited the proliferation rate of two types of hepatocarcinoma cancer cells by more than 45％ compared with everolimus used alone (P<0.01). CONCLUSION The resistance of sirolimus and its derivatives in hepatocellular carcinoma cells may be achieved throngh the feedback-activated PI3K/AKT pathway, and the combination therapy can synergistically inhibit the growth and proliferation of hepatocarcinoma cells.