Search results for: macrophage proliferation

Authors: Kang-Hyun Leem, Myung-Gyou Kim, Hye Kyung Kim

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Velvet antler (VA), the immature antlers of male deer, is traditionally used for thousands of years in Asian countries, such as Korea, China, Taiwan, and Mongolia. It has been considered to improve immune system and physical strength. The goal of this study was to investigate the immunomodulatory effect of deer antler velvet using in vitro system. In the first step, the effects of VA (70% ethanol extract) on the proliferation of splenocytes, bone marrow cell, and macrophages were determined. Next, the effect of VA on the production of nitric oxide and phagocytic activity in macrophage were measured. The results showed that VA treatment increased concanavalin-A stimulated splenocyte, bone marrow cells, and macrophage proliferation in a dose dependent manner. VA at 50 and 100 ug/mL concentrations significantly enhanced the concanavalin-A stimulated splenocyte proliferation by 8.8% and 18.5%, respectively. The proliferation of bone marrow cells, isolated from 5wk-old ICR mice, were increased by 25.2% and 46.5% by 50 and 100 ug/mL VA treatment. RAW 264.7 cell proliferation reached peak value at 50 ug/mL of VA treatment exhibiting 108% of the basal value. Nitric oxide production by RAW 264.7 macrophage cells was slightly reduced by VA treatment but was not statistically significant. Moreover, the phagocytic activity of macrophages was enhanced by VA treatment. These results indicate that VA is effective in immune system.

Keywords: deer antler, splenocyte, bone marrow cells, macrophage proliferation, phagocytosis

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Authors: Hyun Ji Hyun, Hyo Sun Suh, Min Kook Kim, Yong Chan Kwon, Byung-Mu Lee

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Scope: This study is focused on the effect of myristic acid on LPS-induced inflammation in RAW 264.7 macrophage cells. Methods and results: For the experiment, RAW 264.7 mouse macrophage cell line was used. Results showed that treatment with myristic acid can attenuate LPS-induced inflammation. Moreover, myristic acid significantly suppressed expression of inflammatory mediators and down-regulating UVB-induced intracellular ROS generation. Furthermore, myristic acid reduced the expression of NF-κB by inhibiting degradation of IκB-α and ERK, JNK, and p38 pathways by inhibiting phosphorylation in RAW 264.7 macrophage cells. Conclusion: Overall, these data suggest that the myristic acid could reduce LPS-induced inflammation. Acknowledgment: This research was supported by the Ministry of Trade, Industry & Energy(MOTIE), Korea Institute for Advancement of Technology(KIAT) through the Encouragement Program for The Industries of Economic Cooperation Region

Keywords: anti-inflammation, myristic acid, ROS, ultraviolet light

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Authors: Saha Saradindu, Das Payel, Somdeb BoseDasgupta

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Tuberculosis caused by Mycobacteria tuberculosis is a dreadful disease and more so with the advent of extreme and total drug-resistant species. Mycobacterial pathogenesis is an ever-changing paradigm from phagosome maturation block to phagosomal escape into macrophage cytosol and finally acid tolerance and survival inside the lysosome. Mycobacteria are adept at subverting the host immune response by highjacking host cell signaling and secreting virulence factors. One such virulence factor is a ser/thr kinase; Protein kinase G (PknG), which is known to prevent phagosome maturation. The host substrates of PknG, allowing successful pathogenesis still remain an enigma. Hence we carried out a comparative phosphoproteomic screen and identified a number of substrates phosphorylated by PknG. We characterized some of these substrates in vivo and in vitro and observed that PknG mediated phosphorylation of these substrates leads to reduced TNFa production as well as decreased response to TNFa induced macrophage necroptosis, thus enabling mycobacterial survival and proliferation.

Keywords: mycobacteria, Protein kinase G, phosphoproteomics, necroptosis

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Authors: Mansi Srivastava, Uzma Saqib, Adnan Naim, Anjali Roy, Dongfang Liu, Deepak Bhatnagar, Ravinder Ravinder, Mirza S. Baig

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Macrophages polarize to proinflammatory M1 or anti-inflammatory M2 states with distinct physiological functions. This transition within the M1 to M2 phenotypes decides the nature, duration, and severity of an inflammatory response. However, inspite of a substantial understanding of the fate of these phenotypes, the underlying molecular mechanisms are not well understood. We have investigated the role of Neuronal nitric oxide synthase (NOS1) mediated regulation of Activator protein 1 (AP-1) transcription factor in macrophages as a critical effector of macrophage phenotypic change. Activator protein 1 (AP-1) is a group of dimeric transcription factors composed of jun, Fos, and ATF family proteins. We determined that NOS1-derived nitric oxide (NO) facilitate Fos and jun interaction which induces IL12 & IL23 expression. Pharmacological inhibition of NOS1 inhibits Fos and jun interaction but increases ATF2 and Fos dimerization. Switching of Fos and jun dimer to ATF2 and jun dimerization switches phenotype from IL–12high IL-23high IL-10low to IL–12low IL-23lowIL-10high phenotype, respectively. Together, these findings highlight a key role of the TLR4-NOS1-AP1 signaling axis in regulating macrophage polarization.

Keywords: inflammation, macrophage, lipopolysaccharide (LPS), proinflammatory cytokines, activator protein 1 (AP-1), neuronal nitric oxide synthase (NOS1)

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Authors: Z. Chekini, P. Afsharian, F. Ramezanali, A. A. Akhlaghi, R. Aflatoonian

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Macrophage migration inhibitory factor (MIF) is a key pro-inflammatory cytokine that involves in pathophysiological events of endometriosis. We aimed to evaluate the association between mRNA expression levels and polymorphisms of MIF in endometriosis. Seventy endometriosis patients and 70 volunteer fertile women were recruited. RFLP was applied to determine -173G/C polymorphism. ORF polymorphisms and -794(CATT)5-8 were detected by sequencing. Q-PCR was used for expression study of 14 ectopic tissues of patients. Homozygote of CATT5 was observed only in controls. The CATT5/G haplotype related to controls (p=0.094, OR=0.61). Expression level of MIF with -794(CATT)6,7/-173GC was significantly more than the other haplotypes (p=0.00). We identified four SNPs including: +254rs2096525 (p=0.843), +626rs33958703 (p=0.029), +656rs2070766 (p=0.703) and +509rs182012324 (p=1.00). In conclusion, increased repeat of CATT and presence of C allele in promoter of MIF were significantly associated with mRNA level in patients. It seems that +509rs182012324 and +626rs33958703 SNPs were significantly correlated with susceptibility to endometriosis.

Keywords: endometriosis, haplotype, macrophage migration inhibitory factor, polymorphism

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Authors: Zhongyang Sun, Shu Zhang, Manjiang Xie

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Emerging evidence indicates that microRNAs (miRNAs) play important roles in modulating osteoblast function and bone formation. However, the influence of miRNA on osteoblast proliferation and the possible mechanisms underlying remain to be defined. In this study, we aimed to investigate whether miR-103 regulates osteoblast proliferation under simulated microgravity condition through regulating Cav1.2, the primary subunit of L-type voltage sensitive calcium channels (LTCCs). We first investigated the effect of simulated microgravity on osteoblast proliferation and the outcomes clearly demonstrated that the mechanical unloading inhibits MC3T3-E1 osteoblast-like cells proliferation. Using quantitative Real-Time PCR (qRT-PCR), we provided data showing that miR-103 was up-regulated in response to simulated microgravity. In addition, we observed that up-regulation of miR-103 inhibited and down-regulation of miR-103 promoted osteoblast proliferation under simulated microgravity condition. Furthermore, knocking-down or over-expressing miR-103, respectively, up- or down-regulated the level of Cav1.2 expression and LTCCs currents, suggesting that miR-103 acts as an endogenous attenuator of Cav1.2 in osteoblasts under the condition of simulated microgravity. More importantly, we showed that the effect of miR-103 on osteoblast proliferation was diminished in simulated microgravity, when co-transfecting miR-103 mimic or inhibitor with Cav1.2 siRNA. Taken together, our data suggest that miR-103 inhibits osteoblast proliferation mainly through suppression of Cav1.2 expression under simulated microgravity condition. This work may provide a novel mechanism of microgravity-induced detrimental effects on osteoblast, identifying miR-103 as a novel possible therapeutic target in bone remodeling disorders in this mechanical unloading.

Keywords: microRNA, osteoblasts, cell proliferation, Cav1.2, simulated microgravity

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Authors: Cuili Yang, Chengcao Sun, Ruilin Xue, Yongyong Xi, Liang Wang, Dejia Li

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Backgrounds: Previous studies showed that Sirt1 plays an important role in C2C12 myoblast cell proliferation, but the mechanism(s) involved in this process remains unclear. This work was undertaken to determine if Myostatin participates in the regulation of C2C12 proliferation by Sirt1. Methods: We administrated the Sirt1 activator resveratrol, inhibitor Nicotinamide (NAM) and Myostatin inhibitor SB431542 on C2C12 myoblast cells. Cell viability was evaluated by CCK8 assay. The expression of Sirt1 and MyoD were detected by qRT-PCR. Utilizing western blot to determinate the expression of myostatin, P107 and p-P107. Results: Our results showed that resveratrol promoted the proliferation of C2C12 myoblast cells, while NAM suppressed the proliferation of C2C12 myoblast cells; SB431542 promoted the proliferation of C2C12 myoblast cells and attenuated the inhibition effect of NAM on C2C12 myoblast cells proliferation; Resveratrol can significantly increase the expression of Sirt1 and MyoD, decrease the expression of Myostatin, while NAM can significantly down-regulate the expression of Sirt1, MyoD and the phosphorylation of P107(p-P107), but up-regulate the expression of Myostatin and the protein P107; SB431542 can significantly mitigate the effect of NAM on the expression of MyoD, P107, and p-P107. Conclusions: Taken together, these results indicate that Sirt1 promotes the proliferation of C2C12 myoblast cells via Myostatin signaling pathway.

Keywords: Sirt1, C2C12 cells, proliferation, myostatin signaling pathway

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Authors: Forough Ataollahi, Sumit Pramanik, Belinda Pingguan-Murphy, Wan Abu Bakar Bin Wan Abas, Noor Azuan Bin Abu Osman

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Endothelium proliferation is an important process in cardiovascular homeostasis and can be regulated by extracellular environment, as cells can actively sense mechanical environment. In this study, we evaluated endothelial cell proliferation on PDMS/alumina (Al2O3) composites and pure PDMS. The substrates were prepared from pure PDMS and its composites with 5% and 10% Al2O3 at curing temperature 50˚C for 4 h and then characterized by mechanical, structural and morphological analyses. Higher stiffness was found in the composites compared to the pure PDMS substrate. Cell proliferation of the cultured bovine aortic endothelial cells on substrate materials were evaluated via Resazurin assay and 1, 1’-Dioctadecyl-1, 3, 3, 3’, 3’-Tetramethylindocarbocyanine Perchlorate-Acetylated LDL (Dil-Ac-LDL) cell staining, respectively. The results revealed that stiffer substrates promote more endothelial cells proliferation to the less stiff substrates. Therefore, this study firmly hypothesizes that the stiffness elevates endothelial cells proliferation.

Keywords: stiffness, proliferation, bovine aortic endothelial cells, extra cellular matrix, vascular

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Authors: Hager Elsheikh, Matthias Sendler, Juliana Glaubnitz

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In pancreatitis, an inflammatory reaction occurs in the pancreatic secretory cells due to premature activation of proteases, leading to pancreatic self-digestion and necrotic cell death of acinar cells. Acute pancreatitis in patients is characterized by a severe immune reaction that could lead to serious complications, such as organ failure or septic shock, if left untreated. Chronic pancreatitis is a recurrence of episodes of acute pancreatitis resulting in a fibro-inflammatory immune response, in which the type 2 immune response is primarily driven by AAMs in the pancreas. One of the most important signaling pathways for M2 macrophage activation is the IL-4/STAT6 pathway. Pancreatic fibrosis is induced by the hyperactivation of pancreatic stellate cells by dysregulation in the inflammatory response, leading to further damage, autodigestion and possibly necrosis of pancreatic acinar cells. The aim of this research is to investigate the effect of STAT6 knockout in disease severity and development of fibrosis wound healing in the presence of different macrophage populations, regulated by the type 2 immune response, after inducing chronic and/or acute pancreatitis in mice models via cerulean injection. We further investigate the influence of the JAK/STAT6 signaling pathway on the balance of fibrosis and regeneration in STAT6 deficient and wild-type mice. The characterization of resident and recruited macrophages will provide insight into the influence of the JAK/STAT6 signaling pathway on infiltrating cells and, ultimately, tissue fibrosis and disease severity.

Keywords: acute and chronic pancreatitis, tissue regeneration, macrophage polarization, Gastroenterology

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Authors: Ali Oztuna, Meral Sarper, Deniz Torun, Fatma Bayrakdar, Selcuk Kilic, Mehmet Baysallar

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Bacillus anthracis is one of the biological agents most likely to be used in case of bioterrorist attack as well as being the cause of anthrax. The bacterium's major virulence factors are the anthrax toxins and an antiphagocytic polyglutamic capsule. TEM8 (ANTXR1) and CMG2 (ANTXR2) are ubiquitously expressed type I transmembrane proteins, and ANTXR2 is the major receptor for anthrax toxins. MicroRNAs are 21-24 bp small noncoding RNAs that regulate gene expression by base pairing with the 3' UTR (untranslated regions) of their target mRNAs resulting in mRNA degradation and/or translational repression. MicroRNAs contribute to regulation of most biological processes and influence numerous pathological states like infectious disease. In this study, post-exposure (toxins) protective effect of the hsa-miR-124-3p against Bacillus anthracis was examined. In this context, i) THP-1 and U937 cells were differentiated to MΦ macrophage, ii) miRNA transfection efficiencies were evaluated by flow cytometry and qPCR, iii) protection against Bacillus anthracis toxins were investigated by XTT, cAMP ELISA and MEK2 cleavage assays. Acknowledgements: This work was supported by the Scientific and Technological Research Council of Turkey (TUBITAK) under Grant SBAG-218S467.

Keywords: ANTXR2, hsa-miR-124-3p, MΦ macrophage, THP-1, U937

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Authors: Hakim Chayeb

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Macrophages (Mo) play an essential role in host defense against pathogens. These inflammatory cells contain a large group of inducible enzymes such as NO synthase (NOS). This study was conducted to characterize experimentally induced inflammation in vivo by lipopolysaccharides (LPS). LPS is an essential component of the outer membrane of Gram-negative bacteria and a potent inducer of macrophage. Except control rats, all rats received different doses of LPS intra-peritoneally. The involvement of inducible NO synthase (iNOS) and constitutive (cNOS ) in the modulation of the inflammatory response was studied by treating the rats with L-NAME (non-selective NOS inhibitor) or aminoguanidine (AG inhibitor of iNOS). Inhibitors were injected 24 hours before LPS administration. The results showed that esterase activity (a marker of macrophage infiltration) which is induced by LPS is reduced by AG, was potentiated by treatment with L-NAME in tissue homogenates of the liver, kidney and spleen. Meanwhile, the concentrations of nitric oxide (NO) induced by LPS were reduced with AG and are completely inhibited with L-NAME in the tissues studied. NO concentrations and plasma transaminase levels have undergone remarkable increases in rats treated with LPS alone. However, the AG significantly reduced these rates. Our results highlighted the role of NO synthase inhibitors in reducing of inflammatory responses that characterize many infectious diseases.

Keywords: aminoguanidine, esterase, LPS, L-NAME, macrophage, nitric oxide

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Authors: Vijaya Madhuri Devraj, Swarnalatha Guditi, Kiran Kumar Bokara, Gangadhar Taduri

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Cell-based strategies may open therapeutic approaches that promote tolerance through manipulation of macrophages to increase long-term transplant survival rates and minimize side effects of the current immune suppressive regimens. The aim of the present study was, therefore, to test and compare the therapeutic potential of MSC and Tregs on macrophage polarization to develop an alternate cell-based treatment option in kidney transplantation. In the current protocol, macrophages from kidney transplant recipients with graft dysfunction were co-cultured with MSCs and Treg cells with and without cell-cell contact on transwell plates, further to quantitatively assess macrophage polarization in response to MSC and Treg treatment over time, M1 and M2 cell surface markers were used. Additionally, multiple soluble analytes were analyzed in cell supernatant by using bead-based immunoassays. Furthermore, to confirm our findings, gene expression analysis was done. MSCs induced the formation of M2 macrophages more than Tregs when macrophages M0 were cultured in transwell without cell contact. From this, we deduced the mechanism that soluble factors present in the MSCs condition media are involved in skewing of macrophages towards type 2 macrophages; similarly, in co-culture with cell-cell contact, MSCs resulted in more M2 type macrophages than Tregs. And an important finding of this study is the combination of both MSC-Treg showed significantly effective and consistent results in both with and without cell contact setups. Hence, it is suggestive to prefer MSCs over Tregs for secretome-based therapy and a combination of both for either therapy for effective transplantation outcomes. Our findings underline a key role of Tregs and MSCs in promoting macrophage polarization towards anti-inflammatory type. The study has great importance in prolongation of allograft and patient survival without any rejection by cell-based therapy, which induce self-tolerance and controlling infection.

Keywords: graft rejection, graft tolerance, macrophage polarization, mesenchymal stem cells, regulatory T cells, transplant immunology

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Authors: K. Nikovics, D. Riccobono, M. Oger, H. Morin, L. Barbier, T. Poyot, X. Holy, A. Bendahmane, M. Drouet, A. L. Favier

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Muscle regeneration after injury (as irradiation) is of great importance. However, the molecular and cellular mechanisms are still unclear. Cytokines are believed to play fundamental role in the different stages of muscle regeneration. They are secreted by many cell populations, but the predominant producers are macrophages and helper T cells. On the other hand, it has been shown that adipose tissue derived stromal/stem cell (ASC) injection could improve muscle regeneration. Stem cells probably induce the coordinated modulations of gene expression in different macrophage cells. Therefore, we investigated the patterns and timing of changes in gene expression of different cytokines occurring upon stem cells loading. Muscle regeneration was studied in an irradiated muscle of minipig animal model in presence or absence of ASC treatment (irradiated and treated with ASCs, IRR+ASC; irradiated not-treated with ASCs, IRR; and non-irradiated no-IRR). We characterized macrophage populations by immunolabeling in the different conditions. In our study, we found mostly M2 and a few M1 macrophages in the IRR+ASC samples. However, only few M2b macrophages were noticed in the IRR muscles. In addition, we found intensive fibrosis in the IRR samples. With in situ hybridization and immunolabeling, we analyzed the cytokine expression of the different macrophages and we showed that M2d macrophage are the most abundant in the IRR+ASC samples. By in situ hybridization, strong expression of the transforming growth factor β (TGF-β) was observed in the IRR+ASC but very week in the IRR samples. But when we analyzed TGF-β level with immunolabeling the expression was very different: many M2 macrophages showed week expression in IRR+ASC and few cells expressing stronger level in IRR muscles. Therefore, we investigated the MMP expressions in the different muscles. Our data showed that the M2 macrophages of the IRR+ASC muscle expressed MMP2 proteins. Our working hypothesis is that MMP2 expression of the M2 macrophages can decrease fibrosis in the IRR+ASC muscle by capturing TGF-β.

Keywords: adipose tissue derived stromal/stem cell, cytokine, macrophage, muscle regeneration

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Authors: Manh Tin Ho, Phorl Sophors, Ga Young Seo, Young Mee Kim, Youngho Lim, Moonjae Cho

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UV radiation in sunlight is one of the most potential factor induced skin ageing and photocarcinogenesis. UV may induce the melanin production and wrinkle formation. Recently, the natural secondary compounds have been reported that had the beneficial protective effects from UV light. In this study, we investigated the effects of two different compounds, glycitin and 38, on human dermal fibroblast. We first only treated the 38 on melanocyte cell to test the proliferation inhibition of 38 on this cell line. Then, we induced the combination of glycitin and 38 on human dermal fibroblast in 48h and investigate the proliferation, collagen production and the metalloproteinase family expression. The 38 alone could inhibit the proliferation of melanocyte which indicated the reduction of melanin production. The combination of glycitin and 38 truly increased the fibroblast proliferation and even they could recover the UV-induced and H2O2-induced damaged fibroblast proliferation. The co-treatment also promoted the collagen IV expression significantly and accelerated the total collagen secretion. In addition, metalloproteinase (MMPs) family such as MMP1, MMP2, MMP7 was down-regulated in transcriptional level. In conclusion, the combination of glycitin and 38 has induced the fibroblast proliferation even when it was damaged by UV exposure and H2O2, whereas augmented collagen production and inhibited the MMPs caused the wrinkle formation and decreased the melanocyte proliferation, suggested an potential UV-protective therapy.

Keywords: UV radiation, wrinkle, ageing, glycitin, dermal fibroblast

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Authors: Miao Zhang, Limin Liu, Feng Zhi, Panpan Niu, Mengya Yang, Xuemei Zhu, Ying Diao, Jun Wang, Ying Zhao

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Background and Aims: Clinical studies have demonstrated that serum semicarbazide-sensitive amine oxidase (SSAO) activities positively correlate with the progression of atherosclerosis. The aim of the present study is to investigate the effect of SSAO inactivation on the development of atherosclerosis. Methods: Female LDLr knockout (KO) mice were given the Western-type diet for 6 and 9 weeks to induce the formation of early and advanced lesions, and semicarbazide (SCZ, 0.125%) was added into the drinking water to inactivate SSAO in vivo. Results: Despite no impact on plasma total cholesterol levels, abrogation of SSAO by SCZ not only resulted in the enlargement of both early (1.5-fold, p=0.0043) and advanced (1.8-fold, p=0.0013) atherosclerotic lesions, but also led to reduced/increased lesion contents of macrophages/smooth muscle cells (SMCs) (macrophage: ~0.74-fold, p=0.0002(early)/0.0016(advanced); SMC: ~1.55-fold, p=0.0003(early) /0.0001(advanced)), respectively. Moreover, SSAO inactivation inhibited the migration of circulating monocytes into peripheral tissues and reduced the amount of circulating Ly6Chigh monocytes (0.7-fold, p=0.0001), which may account for the reduced macrophage content in lesions. In contrast, the increased number of SMCs in lesions of SCZ-treated mice is attributed to an augmented synthetic vascular SMC phenotype switch as evidenced by the increased proliferation of SMCs and accumulation of collagens in vivo. Conclusion: SSAO inactivation by SCZ promotes the phenotypic switch of SMCs and the development of atherosclerosis. The enzymatic activity of SSAO may thus represent a potential target in the prevention and/or treatment of atherosclerosis.

Keywords: atherosclerosis, phenotype switch of smooth muscle cells, SSAO/VAP-1, semicarbazide

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Authors: Ediati Budi Cahyono, Triana Hertiani, Nauval Arrazy Asawimanda, Wahyu Puji Pratomo

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Background: Doxorubicin is widely used as a chemotherapeutic drug despite having many side effects. It may cause macrophage dysfunction and decreasing proliferation of lymphocyte. Noni (Morinda citrifolia) fruit which has rich of polysaccharide content has potential as antitumor and immunostimulant effect. The isolation of polysaccharide from Noni fruit has been optimized according to four different methods based on macrophage and lymphocyte activities. We found the highest polysaccharide content from one of the four methods isolation. A method of polysaccharide isolation which has the highest immunostimulant effect was used for further observation as co-chemotherapy. The aim of the study: was to evaluate the isolated polysaccharide from the method of choice as co-chemotherapy of doxorubicin for the growth of Vero, He-La, and T47D cell lines in vitro. The method: in vitro growth assay of Vero, He-La, and T47D cell lines was done using MTT-reduction method, and apoptosis test was done by double staining method to evaluate the induction apoptotic effect of the combination. Every group was treated with doxorubicin and isolated polysaccharide from method of choice with 4 variances of concentrations (25 µg/ml, 50 µg/ml, 100 µg/ml and 200 µg/ml) a long with negative control (doxorubicin only) and normal control (without doxorubicin or polysaccharide administration). Results: The combination of polysaccharide fraction in the concentration of 100μg/ml with 2μmol of doxorubicin against He-La and T47D cell lines influenced the highest cytotoxic effect by suppressing cell viability comparing with doxorubicin only. The combination of polysaccharide fraction in the concentration of 100μg/ml with 2μmol of doxorubicin-induced apoptotic effect the He-La cell line comparing with doxorubicin only. The result of the study: it can be concluded that the combination of polysaccharide fraction and doxorubicin effect more selective toward He-La and T47D cell lines than to Vero cell line. It can be suggested isolated polysaccharide from the method of choice has co-chemotherapy activity against doxorubicin.

Keywords: polysaccharide, noni fruit, doxorubicin, cancer cell lines, vero cell line

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Authors: H. Jeries, N. Volkova, C. G. Iglesias, M. Najjar, M. Rosenblat, M. Aviram, T. Hayek

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Background: Synthetic forms of glucocorticoids (e.g., prednisone, prednisolone) are anti-inflammatory drugs which are widely used in clinical practice. The role of glucocorticoids (GCs) in cardiovascular diseases including atherosclerosis is highly controversial, and their impact on macrophage foam cell formation is still unknown. Our aim was to investigate the effects of prednisone or its active metabolite, prednisolone, on macrophage oxidative stress and lipid metabolism using in-vivo, ex-vivo and in-vitro systems. Methods: The in-vivo study included C57BL/6 mice which were intraperitoneally injected with prednisone or prednisolone (5mg/kg) for 4 weeks, followed by lipid metabolism analyses in the mice aorta, and in peritoneal macrophages (MPM). In the ex-vivo study, we analyzed the effect of serum samples obtained from 9 healthy volunteers before or after treatment with oral prednisone (20mg for 5 days), on J774A.1 macrophage atherogenicity. In-vitro studies were conducted using J774A.1 macrophages, human monocyte derived macrophages (HMDM) and fibroblasts. Cells were incubated with increasing concentrations (0-200 ng/ml) of prednisone or prednisolone, followed by determination of cellular oxidative status, triglyceride and cholesterol metabolism. Results: Prednisone or prednisolone treatment resulted in a significant reduction in triglycerides and mainly in cholesterol cellular accumulation in MPM or in J774A.1 macrophages incubated with human serum. Similar resulted were noted in HMDM or in J774A.1 macrophages which were directly incubated with the GCs. These effects were associated with GCs inhibitory effect on triglycerides and cholesterol biosynthesis rates, throughout downregulation of diacylglycerol acyltransferase1 (DGAT1) expression, and of the sterol regulatory element binding protein (SREBP2) and HMGCR expression, respectively. In parallel to prednisone or prednisolone induced reduction in macrophage triglyceride content, paraoxonase 2 (PON2) expression was significantly upregulated. GCs-induced reduction of cellular triglyceride and cholesterol mass was mediated by the GCs receptors on macrophages since the GCs receptor antagonist (RU 486) abolished these effects. In fibroblasts, unlike macrophages, prednisone or prednisolone showed no anti-atherogenic effects. Conclusions: Prednisone or prednisolone are anti-atherogenic since they protected macrophages from lipid accumulation and foam cell formation.

Keywords: atherosclerosis, cholesterol, foam cell, macrophage, prednisone, prednisolone, triglycerides

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Authors: Qing Zhang, Janak L. Pathak, Macro N. Helder, Richard T. Jaspers, Yin Xiao

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Introduction: Nanomaterial-based bone regeneration is greatly influenced by the immune microenvironment. Tissue-engineered nanomaterials mediate the inflammatory response of macrophages to regulate bone regeneration. Silica nanoparticles have been widely used in tissue engineering-related preclinical studies. However, the effect of topological features on the surface of silica nanoparticles on the immune response of macrophages remains unknown. Purposes: The aims of this research are to compare the influences of normal and pollen-like silica nano-surface topography on macrophage immune responses and to obtain insight into their potential regulatory mechanisms. Method: Macrophages (RAW 264.7 cells) were exposed to mesoporous silica nanoparticles with normal morphology (MSNs) and pollen-like morphology (PMSNs). RNA-seq, RT-qPCR, and LSCM were used to assess the changes in expression levels of immune response-related genes and proteins. SEM and TEM were executed to evaluate the contact and adherence of silica nanoparticles by macrophages. For the assessment of the immunomodulation-mediated osteogenic potential, BMSCs were cultured with conditioned medium (CM) from LPS pre-stimulated macrophage cultures treated with MSNs or PMSNs. Osteoimmunomodulatory potential of MSNs and PMSNs in vivo was tested in a mouse cranial bone osteolysis model. Results: The results of the RNA-seq, RT-qPCR, and LSCM assays showed that PMSNs inhibited the expression of pro-inflammatory genes and proteins in macrophages. SEM images showed distinct macrophage membrane surface binding patterns of MSNs and PMSNs. MSNs were more evenly dispersed across the macrophage cell membrane, while PMSNs were aggregated. PMSNs-induced macrophage anti-inflammatory response was associated with upregulation of the cell surface receptor CD28 and inhibition of ERK phosphorylation. TEM images showed that both MSNs and PMSNs could be phagocytosed by macrophages, and inhibiting nanoparticle phagocytosis did not affect the expression of anti-inflammatory genes and proteins. Moreover, PMSNs-induced conditioned medium from macrophages enhanced BMP-2 expression and osteogenic differentiation mBMSCs. Similarly, PMSNs prevented LPS-induced bone resorption via downregulation of inflammatory reaction. Conclusions: PMSNs can promote bone regeneration by modulating osteoimmunological processes through surface topography. The study offers insights into how surface physical contact cues can modulate the regulation of osteoimmunology and provides a basis for the application of nanoparticles with pollen-like morphology to affect immunomodulation in bone tissue engineering and regeneration.

Keywords: physical contact, osteoimmunology, macrophages, silica nanoparticles, surface morphology, membrane receptor, osteogenesis, inflammation

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Authors: Parinaz Tavakoli Zaniani, Dimitrios Balomenos

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Mitochondrial reactive oxygen species (mROS) play a crucial role in macrophage pro-inflammatory activation, although a detailed understanding of the mechanism and kinetics by which mROS drive signaling molecules is still lacking. In general, it is thought that NF-κB activation drives mROS and general ROS production. Here, We performed a detailed kinetic analysis of mROS production during macrophage activation. We found early mROS generation after LPS (lipopolysaccharide) stimulation. Remarkably as early as 5 minutes, mROS signaling promoted initial NF-κB, MAPK activation and pro-inflammatory cytokine production, as established through inhibition or quenching of mROS. On the contrary, NF-κB inhibition had no effect on mROS production. Our findings point to a mechanism by which mROS increase TRAF-6 ubiquitination and, thus NF-κB activity. mROS inhibition reduced LPS-induced lethality in an in vivo septic shock model by controlling pro-inflammatory cytokine production. Overall, our research provides novel insights into the role of mROS as a primary messenger in the pathway of macrophage and as a regulator of inflammatory responses. We found that early mROS production promotes initial NF-κB, and MAPK activation by regulating TRAF-6 ubiquitination and that mROS inhibition can reduce LPS-induced inflammatory cytokines and lethality in a septic shock model. These findings might lead to novel immunotherapeutic strategies targeting early mROS production and control of extreme inflammation in the context of sepsis and other inflammatory diseases.

Keywords: mitochondria, reactive oxygen species, nuclear factor κB, lipopolysaccharide, macrophages

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Authors: Hainan Chen, Jina Qing, Xiao Zhu, Ling Gao, Ampadu O. Jackson, Min Zhang, Kai Yin

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Objective: Editing macrophage activation to dampen inflammatory diseases by promoting the repolarization of inflammatory (M1) macrophages to anti-inflammatory (M2) macrophages is highly associated with mitochondrial respiration. Recent studies have suggested that mitochondrial apolipoprotein A-1 binding protein (APOA1BP) was essential for the cellular metabolite NADHX repair to NADH, which is necessary for the mitochondrial function. The exact role of APOA1BP in the repolarization of M1 to M2, however, is uncertain. Material and method: THP-1-derived macrophages were incubated with LPS (10 ng/ml) or/and IL-4 (100 U/ml) for 24 hours. Biochemical parameters of oxidative phosphorylation and M1/M2 markers were analyzed after overexpression of APOA1BP in cells. Results: Compared with control and IL-4-exposed M2 cells, APOA1BP was downregulated in M1 macrophages. APOA1BP restored the decline in mitochondrial function to improve metabolic and phenotypic reprogramming of M1 to M2 macrophages. Blocking oxidative phosphorylation by oligomycin blunts the effects of APOA1BP on M1 to M2 repolarization. Mechanistically, LPS triggered the hydration of NADH and increased its hydrate NADHX which inhibit cellular NADH dehydrogenases, a key component of electron transport chain for oxidative phosphorylation. APOA1BP decreased the level of NADHX via converting R-NADHX to biologically useful S-NADHX. The mutant of APOA1BP aspartate188, the binding site of NADHX, fail to repair oxidative phosphorylation, thereby preventing repolarization. Conclusions: Restoring mitochondrial function by increasing mitochondrial APOA1BP might be useful to improve the reprogramming of inflammatory macrophages into anti-inflammatory cells to control inflammatory diseases.

Keywords: inflammatory diseases, macrophage repolarization, mitochondrial respiration, apolipoprotein A-1 binding protein, NADHX, NADH

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Authors: Qiaofeng Guo

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Aims: Obesity is often accompanied by insulin resistance (IR) and whole-body inflammation. Aerobic exercise is an effective treatment to improve insulin resistance and inflammation. However, the anti-inflammatory mechanisms of exercise on epididymal and subcutaneous adipose remain to be elucidated. Here, we compared the macrophage polarization between epididymal and subcutaneous adipose after aerobic exercise. Methods: Male C57BL/6 mice were fed a normal diet group or a high-fat diet group for 12 weeks and performed aerobic training on a treadmill at 55%~65% VO₂ max for eight weeks. Food intake, body weight, and fasting blood glucose levels were monitored weekly. The intraperitoneal glucose tolerance test was to evaluate the insulin resistance model. Fat mass, blood lipid profile, serum IL-1β, TNF-α levels, and CD31/CD206 rates were analysed after the intervention. Results: FBG (P<0.01), AUCIPGTT (P<0.01), and HOMA-IR (P<0.01) increased significantly for a high-fat diet and decreased significantly after the exercise. Eight weeks of aerobic exercise attenuated HFD-induced weight gain and glucose intolerance and improved insulin sensitivity. Serum IL-1β, TNF-α, CD11C/CD206 expression in subcutaneous adipose tissue were not changed before and after exercise, but not in epididymal adipose tissue (P<0.01). Conclusion: Insulin resistance is not accompanied by chronic inflammation and M1 polarization of subcutaneous adipose tissue macrophages in high-fat diet mice. Aerobic exercise effectively improved lipid metabolism and insulin sensitivity, which may be closely associated with reduced M1 polarization of epididymal adipose macrophages.

Keywords: aerobic exercise, insulin resistance, chronic inflammation, adipose, macrophage polarization

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Authors: Vagolu S. Krishna, Shan Zheng, Estharla M. Rekha, Luke W. Guddat, Dharmarajan Sriram

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Tuberculosis (TB) remains a major threat to human health. This due to the fact that current drug treatments are less than optimal as well as the rising occurrence of multi drug-resistant and extensively drug-resistant strains of the etiological agent, Mycobacterium tuberculosis (Mt). Given the wide-spread significance of this disease, we have undertaken a design and evaluation program to discover new anti-TB drug leads. Here, our attention is focused on ketol-acid reductoisomerase (KARI), the second enzyme in the branched-chain amino acid biosynthesis pathway. Importantly, this enzyme is present in bacteria but not in humans, making it an attractive proposition for drug discovery. In the present work, we used high-throughput virtual screening to identify seventeen potential inhibitors of KARI using the Birla Institute of Technology and Science in-house database. Compounds were selected based on high docking scores, which were assigned as the result of favourable interactions between the compound and the active site of KARI. The Ki values for two leads, compounds 14 and 16 are 3.71 and 3.06 µM, respectively for Mt KARI. To assess the mode of binding, 100 ns molecular dynamics simulations for these two compounds in association with Mt KARI were performed and showed that the complex was stable with an average RMSD of less than 2.5 Å for all atoms. Compound 16 showed an MIC of 2.06 ± 0.91 µM and a 1.9 fold logarithmic reduction in the growth of Mt in an infected macrophage model. The two compounds exhibited low toxicity against murine macrophage RAW 264.7 cell lines. Thus, both compounds are promising candidates for development as an anti-TB drug leads.

Keywords: ketol-acid reductoisomerase, macrophage, molecular docking and dynamics, tuberculosis

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Authors: Nahid Eskandari, Marjan Ghagozolo, Ehsan Almasi

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Introduction: Silymarin, as a polyphenolic flavonoid derived from milk thistle (Silybum marianum), is known to have antioxidant, immunomodulatory, antiproliferative, antifibrotic, and antiviral effects. The goal of this study was to determine immunosuppressive effect of Silymarin on proliferation and apoptosis of human T cells in comparison with Rapamycin and FK506. Methods: Peripheral Blood Mononuclear Cells (PBMCs) from healthy individuals were activated with Con A (5µg/ml) and then treated with Silymarin, Rapamycin and FK506 in various concentrations (0.001, 0.01, 0.1, 1, 10,100 and 200M) for 5 days. PBMCs were examined for proliferation using CFSE assay and the concentration that inhibited 50% of the cell proliferation (IC50) was determined for each treatment. For apoptosis assay using flow cytometry, PBMCs were activated with Con A and treated with IC50 dose of Silymarin, Rapamycin and FK506 for 5 days, then cell apoptosis was analysed by FITC-annexin V/PI staining and flow cytometry. The effects of Silymarin, Rapamycin and FK506 on the activation of PARP (poly ADP ribose polymerase) pathway in PBMCs stimulated with Con A and treated with IC50 dose of drugs for 5 days evaluated using the PathScan cleaved PARP sandwich ELISA kit. Results: This study showed that Silymarin had the ability to inhibit T cell proliferation in vitro. Moreover, our results indicated that 100 μM (P < 0.001) and 200 μM (P < 0.001) of Silymarin has more inhibitory effect on T cells proliferation than FK506 and Rapamycin. Our data showed that the effective doses (IC50) of Silymarin, FK506 and Rapamycin were 3×10-5 µM, 10-8 µM and 10-6 µM respectively. Data showed that the inhibitory effect of Silymarin, FK506 and Rapamycin on T cell proliferation was not due to cytotoxicity and none of these drugs at IC50 concentration had not affected the level of cleaved PARP. Conclusion: Silymarin could be a good candidate for immunosuppressive therapy for certain medical conditions with superior efficacy and lesser toxicity in comparison with other immunosuppressive drugs.

Keywords: silymarin, immunosuppressive effect, rapamycin, immunology

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Authors: Uche A. Nnawulezi

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This paper analyzes the advancing nature of nuclear terrorism and proliferation in the global environment and its attendant impacts. It analyzes discourse and practice with respect to the general prohibition on the utilization of fissionable radioactive materials. Thus, there has been a few ideological, reasonable and academic recommendations of policies aimed at eliminating nuclear weapons which its ultimate nightmare has remained an assault including nuclear explosion in densely populated urban areas. Likewise, this paper concentrates on safety measures aimed at preventing nuclear assaults which should not just concentrate on endeavors to prevent terrorists from exploding nuclear gadgets but should be more concerned on endeavors aimed at preventing the acquisition of nuclear weapons in the first place. The author of this paper has pointed out that the non-proliferation treaty should be vigorously supported as well as the Comprehensive Test Ban Treaty brought into force. This paper depended unequivocally on secondary sources, for example, textbooks, journals, articles, and periodicals. It concludes that the fundamental proposals made in this paper if completely used shall remain a cornerstone of efforts made in preventing the spread of nuclear weapons. At last, the only way is to eliminate stockpiles of nuclear weapons in the world or else the likelihood of nuclear terrorism remains a nightmare.

Keywords: nuclear, terrorism, proliferation, global environment

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Authors: Mumuni Sumaila, Viness Pillay, Yahya E. Choonara, Pradeep Kumar, Pierre P. Kondiah

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Tuberculosis (TB) remains a serious challenge to public health globally, despite every effort put together to curb the disease. Current TB therapeutics available have proven to be inefficient due to a multitude of drawbacks that range from serious adverse effects/drug toxicity to inconsistent bioavailability, which ultimately contributes to the emergence of drug-resistant TB. An effective ‘cargo’ system designed to cleverly deliver therapeutic doses of anti-TB drugs to infection sites and in a sustained-release manner may provide a better therapeutic choice towards winning the war against TB. In the current study, we investigated mannose-functionalized lipopolysaccharide hybrid nanoparticles for safety and efficacy towards macrophage-targeted simultaneous delivery of the two first-line anti-TB drugs, rifampicin (RF) and isoniazid (IS). RF-IS-loaded lipopolysaccharide hybrid nanoparticles were fabricated using the solvent injection technique (SIT), incorporating soy lecithin (SL) and low molecular weight chitosan (CS) as the lipid and polysaccharide components, respectively. Surface-functionalized nanoparticles were obtained through the reaction of the aldehyde group of mannose with free amine functionality present at the surface of the nanoparticles. The functionalized nanocarriers were spherical with average particle size and surface charge of 107.83 nm and +21.77 mV, respectively, and entrapment efficiencies (EE) were 53.52% and 69.80% for RF and IS, respectively. FTIR spectrum revealed high-intensity bands between 1663 cm⁻¹ and 1408 cm⁻¹ wavenumbers (absent in non-functionalized nanoparticles), which could be attributed to the C=N stretching vibration produced by the formation of Schiff’s base (–N=CH–) during the mannosylation reaction. In vitro release studies showed a sustained-release profile for RF and IS, with less than half of the total payload released over a 48-hour period. The nanocarriers were biocompatible and safe, with more than 80% cell viability achieved when incubated with RAW 264.7 cells at concentrations 30 to 500 μg/mL over a 24-hour period. Cellular uptake studies (after a 24-hour incubation period with the murine macrophage cells, RAW 264.7) revealed a 13- and a 9-fold increase in intracellular accumulation of RF and IS, respectively, when compared with the unformulated RF+IS solution. A 6- and a 3-fold increase in intracellular accumulation of RF and IS, respectively, were observed when compared with the non-functionalized nanoparticles. Furthermore, fluorescent microscopy images showed nanoparticle internalization and accumulation within the RAW 264.7 cells, which was more significant in the mannose-functionalized system compared to the non-functionalized nanoparticles. The overall results suggested that the fabricated mannose-functionalized lipopolysaccharide nanoparticles are a safe and promising platform for macrophage-targeted delivery of anti-TB therapeutics. However, in vivo pharmacokinetic/pharmacodynamics studies are required to further substantiate the therapeutic efficacy of the nanosystem.

Keywords: anti-tuberculosis therapeutics, hybrid nanosystem, lipopolysaccharide nanoparticles, macrophage-targeted delivery

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Authors: Isha Ranadive, Sonam Patel, Suresh Balakrishnan

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It has been observed that in lizards wound can be healed by either a scar free mechanism or by scarring. The animal model used to study both these healing processes was Northern House Gecko. In lizard, the tail when amputated heals by scar free mechanism which allows it to regenerate, the same is not seen when the limb is amputated. Proliferation, apoptosis, and angiogenesis are the main events which succeed an injury. We observed that proliferation of the cells beneath the wound epidermis was much higher in case of wound healing in tail. This could be because after the wound gets covered by the epithelium, it enters in to a cross-talk with the underlying mesenchyme to recruit a pool of blastemal cells which proliferate and later differentiate to form the lost part through epimorphic regeneration. This was substantiated by mRNA expression levels of various FGFs which facilitate the cross-talk and also by PCNA which is a marker for proliferation. Western blot result reaffirms the same notion. However, in case of the limb, the rate of apoptosis was more than proliferation as there are a lot of debris that needs to be removed. We came to this conclusion as we observed that p53 the apoptotic gene was highly upregulated in case of the scarred tissue. Further, we confirmed this result by checking the anti-apoptotic gene bcl2 and found it to be significantly down-regulated. As we noticed heightened proliferation in the case of scar-free wound healing in tail, angiogenesis was targeted for the study. This is because, when the cells are proliferating they require constant supply of blood and hence neo-vascularization is inevitable. It was observed that the marker of angiogenesis, VEGF, was expressed more during wound healing as compared to the resting stage of tail. Moreover, a high up-regulation was seen in KDR, a receptor of VEGF. Thus, this study reveals how proliferation, apoptosis, and angiogenesis play a key role in the scar-free as well as scarred wound healing.

Keywords: epimorphic regeneration, injury, northern house gecko, wound healing

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Authors: Magdalena Rusady Goey, Gordon Strathdee, Neil Perkins

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Background: Acute lymphoblastic leukaemia (ALL) is the most frequent type of childhood malignancy, accounting for 25% of all cases. TWIST2, a basic helix-loop-helix transcription factor, has been implicated in ALL development. Prior studies found that TWIST2 undergoes epigenetic silencing in more than 50% cases of ALL through promoter hypermethylation and suggested that re-expression of TWIST2 may inhibit cell growth/survival of leukaemia cell lines. TWIST2 has also been implicated as a regulator of NF-kappaB activity, which is constitutively active in leukaemia. Here, we use a lentiviral transductions system to confirm the importance of TWIST2 in controlling leukaemia cell growth and to investigate whether this is achieved through altered regulation of NF-kappaB activity. Method: Re-expression of TWIST2 in leukaemia cell lines was achieved using lentiviral-based transduction. The lentiviral vector also expresses enhanced green fluorescent protein (eGFP), allowing transduced cells to be tracked using flow cytometry. Analysis of apoptosis and cell proliferation were done using annexinV and VPD450 staining, respectively. Result and Discussion: TWIST2-expressing cells were rapidly depleted from a mixed population in ALL cell lines (NALM6 and Reh), indicating that TWIST2 inhibited cell growth/survival of ALL cells. In contrast, myeloid cell lines (HL60 and K562) were comparatively insensitive to TWIST2 re-expression. Analysis of apoptosis and cell proliferation found no significant induction of apoptosis, but did find a rapid induction of proliferation arrest in TWIST2-expressing Reh and NALM6 cells. Initial experiment with NF-kappaB inhibitor demonstrated that inhibition of NF-kappaB has similar impact on cell proliferation in the ALL cell lines, suggesting that TWITST2 may induce cell proliferation arrest through inhibition of NF-kappaB. Conclusion: The results of this study suggest that epigenetic inactivation of TWIST2 in primary ALL leads to increased proliferation, potentially by altering the regulation of NF-kappaB.

Keywords: leukaemia, acute lymphoblastic leukaemia, NF-kappaB, TWIST2, lentivirus

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Authors: Marasri Junsi, Sunisa Siripongvutikorn, Chutha Takahashi Yupanqui, Worrapong Usawakesmanee

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Thunbergia laurifolia has been used for folklore medicine purposes and consumed in the form of herbal tea in Thailand since ancient times. To evaluate the bioactive compounds of aqueous leave extract possessed antioxidant and anti-inflammatory activities. The antioxidant activities were examined by total extractable phenolic content (TPC), total extractable flavonoid content (TFC), ABTS radical scavenging, DPPH radical scavenging, FRAP reducing antioxidant power expressed as mg of gallic acid trolox and caffeic acid for the equivalents. Results indicated that the extract had high TPC and antioxidant activities. In addition, the HPLC-DAD analysis of phenolics and flavonoids indicated the presence of caffeic acid and rutin as bioactive compounds. Exposure of cells with the extract using nitric oxide (NO) production in RAW 264.7 murine macrophage cell line induced by lipopolysaccharide (LPS) was significantly reduced NO production and increased cell proliferation. The obtained results demonstrated that the extract contains a high potential to be used as anti-inflammatory and antioxidant substances.

Keywords: Thunbergia laurifolia, anti-inflammatory, antioxidant activities, RAW264.7

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Authors: Shangerganesh Lingeshwaran

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In this presentation, we have performed numerical simulations for a reaction-diffusion equation with various nonlinear density-dependent diffusion operators and proliferation functions. The mathematical model represented by parabolic partial differential equation is considered to study the invasion of gliomas (the most common type of brain tumors) and to describe the growth of cancer cells and response to their treatment. The unknown quantity of the given reaction-diffusion equation is the density of cancer cells and the mathematical model based on the proliferation and migration of glioma cells. A standard Galerkin finite element method is used to perform the numerical simulations of the given model. Finally, important observations on the each of nonlinear diffusion functions and proliferation functions are presented with the help of computational results.

Keywords: glioma invasion, nonlinear diffusion, reaction-diffusion, finite eleament method

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Authors: Nehir Nebioglu

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Chemotherapy is widely used for the treatment of cancer. Doxorubicin is an anti-cancer chemotherapy drug that is classified as an anthracycline antibiotic. Antitumor antibiotics consist of natural products produced by species of the soil fungus Streptomyces. These drugs act in multiple phases of the cell cycle and are known cell-cycle specific. Although DOX is a precious clinical antineoplastic agent, resistance is also a problem that limits its utility besides cardiotoxicity problem. The drug resistance of cancer cells results from multiple factors including individual variation, genetic heterogeneity within a tumor, and cellular evolution. The mechanism of resistance is thought to involve, in particular, ABCB1 (MDR1, Pgp) and ABCC1 (MRP1) as well as other transporters. Several studies on DOX-resistant cell lines have shown that resistance can be overcome by an inhibition of ABCB1, ABCC1, and ABCC2. This study attempts to understand the effects of different concentration levels of glucose treatment and starvation on the proliferation of Doxorubicin resistant cancer cells lines. To understand the effect of starvation, K562/Dox and K562 cell lines were treated with 0, 5 nM, 50 nM, 500 nM, 5 uM and 50 uM Dox concentrations in both starvation and normal medium conditions. In addition to this, to interpret the effect of glucose treatment, different concentrations (0, 1 mM, 5 mM, 25 mM) of glucose were applied to Dox-treated (with 0, 5 nM, 50 nM, 500 nM, 5 uM and 50 uM) K562/Dox and K652 cell lines. All results show significant decreasing in the cell count of K562/Dox, when cells were starved. However, while proliferation of K562/Dox lines decrease is associated with the increasingly applied Dox concentration, K562/Dox starved ones remain at the same proliferation level. Thus, the results imply that an amount of K562/Dox lines gain starvation resistance and remain resistant. Furthermore, for K562/Dox, there is no clear effect of glucose treatment in terms of cell proliferation. In the presence of a moderate level of glucose (5 mM), proliferation increases compared to other concentration of glucose for each different Dox application. On the other hand, a significant increase in cell proliferation in moderate level of glucose is only observed in 5 uM Dox concentration. The moderate concentration level of Dox can be examined in further studies. For the high amount of glucose (25 mM), cell proliferation levels are lower than moderate glucose application. The reason could be high amount of glucose may not be absorbable by cells. Also, in the presence of low amount of glucose, proliferation is decreasing in an orderly manner of increase in Dox concentration. This situation can be explained by the glucose depletion -Warburg effect- in the literature.

Keywords: drug resistance, cancer cells, chemotherapy, doxorubicin

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