Autophagy modulation effect on homotypic transfer of intracellular components via tunneling nanotubes in mesenchymal stem cells.

BACKGROUND: Recent studies have proved the role of autophagy in mesenchymal stem cell (MSCs) function and regenerative properties. How and by which mechanism autophagy modulation can affect the juxtacrine interaction of MSCs should be addressed. Here, the role of autophagy was investigated in the formation of tunneling nanotubes (TNTs) and homotypic mitochondrial donation. METHODS: MSCs were incubated with 15 µM Metformin (Met) and/or 3 µM 3-methyladenine (3-MA) for 48 h. The formation of TNTs was assessed using bright-field and SEM images. The mitochondria density and ΔΨ values were monitored using flow cytometry analysis. Using RT-PCR and protein array, the close interaction and shared mediators between autophagy, apoptosis, and Wnt signaling pathways were also monitored. The total fatty acid profile was assessed using gas chromatography. RESULT: Data indicated the increase of TNT length and number, along with other cell projections after the induction of autophagy while these features were blunted in 3-MA-treated MSCs (p < 0.05). Western blotting revealed the significant reduction of Rab8 and p-FAK in 3-MA-treated MSCs (p < 0.05), indicating the inhibition of TNT assembly and vesicle transport. Likewise, the stimulation of autophagy increased autophagic flux and mitochondrial membrane integrity compared to 3-MA-treated MSCs. Despite these findings, protein levels of mitochondrial membrane Miro1 and 2 were unchanged after autophagy inhibition/stimulation (p > 0.05). We found that the inhibition/stimulation of autophagy can affect the protein, and transcription levels of several mediators related to Wnt and apoptosis signaling pathways involved in different cell bioactivities. Data confirmed the profound increase of mono and polyunsaturated/saturated fatty acid ratio in MSCs exposed to autophagy stimulator. CONCLUSIONS: In summary, autophagy modulation could affect TNT formation which is required for homotypic mitochondrial donation. Thus, the modulation of autophagy creates a promising perspective to increase the efficiency of cell-based therapies.

Tumoricidal properties of thymoquinone on human colorectal adenocarcinoma cells via the modulation of autophagy.

Colorectal cancer (CRC) is deadly anaplastic changes in the gastrointestinal tract with high-rate mortality. In recent years, the application of phytocompounds has been extended along with different therapeutic protocols. Here, we monitored the effects of Thymoquinone (TQ) on autophagy via mitochondrial function after modulation of the Wnt/ß-catenin signaling pathway.Human colorectal adenocarcinoma HT-29 cells were treated with TQ (60 µM) and 15 µM Wnt3a inhibitor (LGK974) for 48 h. The survival rate was evaluated using an MTT assay. The expression of Wnt-related factors (c-Myc, and Axin), angiogenesis (VE-Cadherin), and mitophagy-related factors (PINK1, OPTN) was assessed using real-time PCR assay. Protein levels of autophagy factors (Beclin-1, LC3, and P62) were monitored using western blotting. Using flow cytometry analysis, the intracellular accumulation of Rhodamine 123 was evaluated. The migration properties were analyzed using a scratch wound healing assay.Data indicated that TQ can reduce the viability of HT-29 cells compared to the control cells (p < 0.05). The expression of VE-Cadherin was inhibited while the expression of PINK1 was induced in treated cells (p < 0.05). Both LGK974 and TQ-treated cells exhibited activation of autophagy flux (Beclin-1↑, LC3II/I↑, and p62↓) compared to the control group (p < 0.05). TQ can increase intracellular accumulation of Rhodamine 123, indicating the inhibition of efflux mechanisms in cancer cells. Along with these changes, the migration of cells was also reduced (p < 0.05).TQ is a potential phytocompound to alter the dynamic growth of human colorectal HT-29 cells via the modulation of autophagy, and mitophagy-related mechanisms.

Mitochondria-loaded alginate-based hydrogel accelerated angiogenesis in a rat model of acute myocardial infarction.

Here, mitochondria were isolated from mesenchymal stem cells (MSCs) after being treated with mitochondria-stimulating substrates, 50 µM metformin (Met), and 40 µM dichloroacetic acid (DCA). The isolated mitochondria (2 × 107 particles) were characterized and encapsulated inside 100 µl hydrogel composed of alginate (3 % w/v; Alg)/gelatin (Gel; 1 % w/v) enriched with 1 µM pyrrole (Pyr) solidified in the presence of 0.2 M FeCl3. The physicochemical properties and cytocompatibility of prepared hydrogels were assessed using FTIR, swelling, biodegradation, porosity assays, and scanning electron microscopy (SEM). The mitochondria-bearing hydrogel was injected into the ischemic area of rat hearts. FTIR absorption bands represented that the addition of FeCl3 led to polypyrrole (PPy) formation, polysaccharide oxidation, and interaction between Alg and Gel. SEM images exhibited porous structure and the size of pores was reduced in Alg/Gel + PPy group compared to Alg + PPy hydrogel. Based on the data, both Alg + PPy and Alg/Gel + PPy hydrogels can preserve the integrity and morphology of loaded mitochondria. It was noted that Alg/Gel + PPy hydrogel possessed a higher swelling ratio, degradation, and porosity compared to Alg + PPy group. Data confirmed that Alg/Gel + PPy hydrogel containing 1 µM Pyr yielded the highest survival rate compared to groups with 2 and 4 µM Pyr (p < 0.05). Injection of mitochondria-loaded Alg/Gel + PPy hydrogel yielded significant restoration of left ventricle thickness compared to the infarction, mitochondria, and Alg/Gel + PPy hydrogel groups 14 days post-injection (p < 0.05). Histological analyses revealed a significant increase of vWF+ capillaries and α-SMA+ arterioles in the mitochondria-loaded Alg/Gel + PPy hydrogel group (p < 0.05). Immunofluorescence imaging revealed the ability of rat cardiomyocytes to uptake mitochondria alone or after being loaded into Alg/Gel + PPy hydrogel. These effects were evident in the Alg/Gel + PPy group. Taken together, electroconductive Alg-based hydrogels are suitable platforms for the transplantation of cells and organelles and the regeneration of ischemic heart changes.

Angiogenic activity of mitochondria; beyond the sole bioenergetic organelle.

Angiogenesis is a complex process that involves the expansion of the pre-existing vascular plexus to enhance oxygen and nutrient delivery and is stimulated by various factors, including hypoxia. Since the process of angiogenesis requires a lot of energy, mitochondria play an important role in regulating and promoting this phenomenon. Besides their roles as an oxidative metabolism base, mitochondria are potential bioenergetics organelles to maintain cellular homeostasis via sensing alteration in oxygen levels. Under hypoxic conditions, mitochondria can regulate angiogenesis through different factors. It has been indicated that unidirectional and bidirectional exchange of mitochondria or their related byproducts between the cells is orchestrated via different intercellular mechanisms such as tunneling nanotubes, extracellular vesicles, and gap junctions to maintain the cell homeostasis. Even though, the transfer of mitochondria is one possible mechanism by which cells can promote and regulate the process of angiogenesis under reperfusion/ischemia injury. Despite the existence of a close relationship between mitochondrial donation and angiogenic response in different cell types, the precise molecular mechanisms associated with this phenomenon remain unclear. Here, we aimed to highlight the possible role of mitochondria concerning angiogenesis, especially the role of mitochondrial transport and the possible relation of this transfer with autophagy, the housekeeping phenomenon of cells, and angiogenesis.

Vandetanib alters the tumoricidal capacity of human breast cancer stem cells via inhibiting vasculogenic capacity.

Introduction: The inhibition of vascularization into tumor stroma as well as dynamic cell growth is the center of attention. Here, we aimed to examine the role of vandetanib on angiogenesis capacity of breast cancer stem cell (CSCs). Methods: MDA-MB-231 cells were exposed to different doses of vandetanib and survival rate was monitored. Stimulatory effects of vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), and epidermal growth factor (EGF) were evaluated in vandetanib-treated MDA-MB-231 cells. In vitro tubulogenesis capacity was studied on the Matrigel surface. The synergistic effects of vandetanib on cell survival were also assessed after PI3K and/or Wnt3a inhibition. Vascular endothelial (VE)-cadherin, matrix metalloproteinase-2 (MMP-2), -9, Wnt3a, and p-Akt/Akt ratio were measured using western blotting. Results: Vandetanib reduced survival rate in a dose-dependent manner (P<0.05). Proliferative effects associated with VEGF, FGF, and EGF were blunted in these cells pre-exposed to vandetanib (P<0.05). The microcirculation pattern's triple-negative breast cancer (TNBC) was suppressed by 1, 5 µM of vandetanib (P<0.05). Hence 1, 5 µM of vandetanib potentially decreased the population of CD24- cells. 1 and 5 µM of vandetanib inhibited cell proliferation by blocking PI3K and Wnt3a pathways and decreased the p-Akt/Akt ratio, Wnta3 protein levels (P<0.05). 1 and 5 µM vandetanib combined with PI3K inhibitor diminished metastatic markers including, MMP-2, and MMP-9. The concurrent treatment (PI3K, inhibitor+ 1, 5 µM vandetanib) also considerably reduced epithelial-mesenchymal transition (EMT) markers such as VE-cadherin (P<0.05). Conclusion: Vandetanib suppressed vasculogenic mimicry (VM) networking through blunting stemness properties, coincided with suppression of VE-cadherin in CSCs.

Polyurethane-based nanofibrous mat containing porphyrin with photosensitivity and bactericidal properties can promote cutaneous tissue healing in rats.

The regeneration of cutaneous tissue is one of the most challenging issues in human regenerative medicine. To date, several studies have been done to promote cutaneous tissue healing with minimum side effects. The healing potential of polyurethane (PU)/Poly (caprolactone)-poly (ethylene glycol)-poly (caprolactone) (PCEC)/chitosan (CS) (PCS) nanofibrous mat with cationic photosensitizer meso tetrakis (N-methyl pyridinium-4-yl) porphyrin tetratosylate salt (TMP) was examined. The CS tripolyphosphate nanoparticles (CSNPs) were prepared and loaded by TMP to provide an efficient drug release system (TMPNPs) for delivery of TMP to promote wound healing. In in vitro setting, parameters such as bactericidal effects, cytocompatibility, and hemolytic effects were examined. The healing potential of prepared nanofibrous mats was investigated in a rat model of full-thickness cutaneous injury. PCS/TMP/TMPNPs nanofibers can efficiently release porphyrin in the aqueous phase. The addition of TMPNPs and CS to the PU backbone increased the hydrophilicity, degradation, and reduced mechanical properties. The culture of human fetal foreskin fibroblasts (HFFF2) on PCS/TMP/TMPNPs scaffold led to an increased survival rate and morphological adaptation analyzed by MTT and SEM images. Irradiation with a red laser (635 nm, 3 J/cm2) for the 30 s reduced viability of S. aureus and E. Coli bacteria plated on PCS/TMP and PCS/TMP/TMPNPs nanofibrous mats compared to PU/PCEC (PC) and PU/PCEC/CS (PCS) groups, indicating prominent antibacterial effects of PCS/TMP and PCS/TMP/TMPNPs nanofibrous (p < 0.05). Data indicated that PCS/TMP/TMPNPs mat enhanced healing of the full-thickness excisional wound in a rat model by the reduction of inflammatory response and fibrotic changes compared to the PC, and PCS groups (p < 0.05). Immunofluorescence imaging indicated that levels of Desmoglein were increased in rats that received PCS/TMP/TMPNPs compared to the other groups. It is found that a PU-based nanofibrous mat is an appropriate scaffold to accelerate the healing of injured skin.

The Beneficial and Adverse Effects of Autophagic Response to Caloric Restriction and Fasting.

Each cell is equipped with a conserved housekeeping mechanism, known as autophagy, to recycle exhausted materials and dispose of injured organelles via lysosomal degradation. Autophagy is an early-stage cellular response to stress stimuli in both physiological and pathological situations. It is thought that the promotion of autophagy flux prevents host cells from subsequent injuries by removing damaged organelles and misfolded proteins. As a correlate, the modulation of autophagy is suggested as a therapeutic approach in diverse pathological conditions. Accumulated evidence suggests that intermittent fasting or calorie restriction can lead to the induction of adaptive autophagy and increase longevity of eukaryotic cells. However, prolonged calorie restriction with excessive autophagy response is harmful and can stimulate a type II autophagic cell death. Despite the existence of a close relationship between calorie deprivation and autophagic response in different cell types, the precise molecular mechanisms associated with this phenomenon remain unclear. Here, we aimed to highlight the possible effects of prolonged and short-term calorie restriction on autophagic response and cell homeostasis.

Melatonin blunted the angiogenic activity in 3D colon cancer tumoroids by the reduction of endocan.

BACKGROUND: Complexity and heterogeneity of the tumor niche are closely associated with the failure of therapeutic protocols. Unfortunately, most data have been obtained from conventional 2D culture systems which are not completely comparable to in vivo microenvironments. Reconstructed 3D cultures composed of multiple cells are valid cell-based tumor models to recapitulate in vivo-like interaction between the cancer cells and stromal cells and the oncostatic properties of therapeutics. Here, we aimed to assess the tumoricidal properties of melatonin on close-to-real colon cancer tumoroids in in vitro conditions. METHODS: Using the hanging drop method, colon cancer tumoroids composed of three cell lines, including adenocarcinoma HT-29 cells, fibroblasts (HFFF2), and endothelial cells (HUVECs) at a ratio of 2: 1: 1, respectively were developed using 2.5% methylcellulose. Tumoroids were exposed to different concentrations of melatonin, from 0.005 to 0.8 mM and 4 to 10 mM, for 48 h. The survival rate was measured by MTT and LDH leakage assays. Protein levels of endocan and VEGF were assessed using western blotting. Using histological examination (H & E) staining, the integrity of cells within the tumoroid parenchyma was monitored. RESULTS: Despite the reduction of viability rate in lower doses, the structure of tumoroids remained unchanged. In contrast, treatment of tumoroids with higher doses of melatonin, 4 and 10 mM, led to disaggregation of cells and reduction of tumoroid diameter compared to the non-treated control tumoroids (p < 0.05). By increasing melatonin concentration from 4 to 10 mM, the number of necrotic cells increased. Data showed the significant suppression of endocan in melatonin-treated tumoroids related to the non-treated controls (p < 0.05). According to our data, melatonin in higher doses did not alter protein levels of VEGF (p > 0.05). CONCLUSIONS: Melatonin can exert its tumoricidal properties on colon cancer tumoroids via the reduction of tumor cell viability and inhibition of the specific pro-angiogenesis factor.

Interaction of lncRNAs with mTOR in colorectal cancer: a systematic review.

Colorectal cancer (CRC) is the third most widespread cancer and the fourth leading lethal disease among different societies. It is thought that CRC accounts for about 10% of all newly diagnosed cancer cases with high-rate mortality. lncRNAs, belonging to non-coding RNAs, are involved in varied cell bioactivities. Emerging data have confirmed a significant alteration in lncRNA transcription under anaplastic conditions. This systematic review aimed to assess the possible influence of abnormal mTOR-associated lncRNAs in the tumorigenesis of colorectal tissue. In this study, the PRISMA guideline was utilized based on the systematic investigation of published articles from seven databases. Of the 200 entries, 24 articles met inclusion criteria and were used for subsequent analyses. Of note, 23 lncRNAs were prioritized in association with the mTOR signaling pathway with up-regulation (79.16%) and down-regulation (20.84%) trends. Based on the obtained data, mTOR can be stimulated or inhibited during CRC by the alteration of several lncRNAs. Determining the dynamic activity of mTOR and relevant signaling pathways via lncRNAs can help us progress novel molecular therapeutics and medications.

Exosomal transmission of viruses, a two-edged biological sword.

As a common belief, most viruses can egress from the host cells as single particles and transmit to uninfected cells. Emerging data have revealed en bloc viral transmission as lipid bilayer-cloaked particles via extracellular vesicles especially exosomes (Exo). The supporting membrane can be originated from multivesicular bodies during intra-luminal vesicle formation and autophagic response. Exo are nano-sized particles, ranging from 40-200 nm, with the ability to harbor several types of signaling molecules from donor to acceptor cells in a paracrine manner, resulting in the modulation of specific signaling reactions in target cells. The phenomenon of Exo biogenesis consists of multiple and complex biological steps with the participation of diverse constituents and molecular pathways. Due to similarities between Exo biogenesis and virus replication and the existence of shared pathways, it is thought that viruses can hijack the Exo biogenesis machinery to spread and evade immune cells. To this end, Exo can transmit complete virions (as single units or aggregates), separate viral components, and naked genetic materials. The current review article aims to scrutinize challenges and opportunities related to the exosomal delivery of viruses in terms of viral infections and public health. Video Abstract.

Hyper-Branched Cationic Cyclodextrin Polymers for Improving Plasmid Transfection in 2D and 3D Spheroid Cells.

In this article, we used monolayer two dimensional (2D) and 3D multicellular spheroid models to improve our understanding of the gene delivery process of a new modified cationic hyper-branched cyclodextrin-based polymer (Ppoly)-loaded plasmid encoding Enhanced Green Fluorescent Protein (EGFP). A comparison between the cytotoxicity effect and transfection efficiency of the plasmid DNA (pDNA)-loaded Ppoly system in 2D and 3D spheroid cells determined that the transfection efficiency and cytotoxicity of Ppoly-pDNA nanocomplexes were lower in 3D spheroids than in 2D monolayer cells. Furthermore, histopathology visualization of Ppoly-pDNA complex cellular uptake in 3D spheroids demonstrated that Ppoly penetrated into the inner layers. This study indicated that the Ppoly, as a non-viral gene delivery system in complex with pDNA, is hemocompatible, non-toxic, high in encapsulation efficiency, and has good transfection efficiency in both 2D and 3D cell cultures compared to free pDNA and lipofectamine (as the control).

In vitro exosomal transfer of Nrf2 led to the oxaliplatin resistance in human colorectal cancer LS174T cells.

Chemotherapy resistance is a serious pitfall in the treatment of colon cancers (CCs). Previous studies have found that exosomes (Exo) play a pivotal role in tumor drug resistance via the transfer of proteins and genetic materials to the acceptor cells. To date, the mechanisms orchestrating Exo-derived resistance in cancer cells have been the center of attention. Herein, we aimed to evaluate the role of exosomal nuclear factor erythroid 2-related factor 2 (Nrf2) on oxaliplatin (1-OHP) resistance in human colorectal cancer LS174T cells in vitro. To this end, exosomal-Nrf2-mediated 1-OHP resistance was examined using different assays. Exo was isolated from resistant LS174T cells (LS174T/R) and added to the culture medium of sensitive LS174T cells (LS174T/S). According to our data, LS174T/S cells successfully adsorbed PKH26-Exo driven from LS174T/R cells. Western blotting showed an increased Nrf2 level in Exo isolated from LS174T/R cells compared to LS174T/S cell-derived Exo (p < .05). The incubation of LS174T/S cells with LS174T/R-derived Exo increased half-maximal inhibitory concentration values in response to treatment with 1-OHP (p < .05). Besides this, the apoptotic changes were diminished in LS174T/S cells after incubation with LS174T/R-derived Exo. Of note, the exposure of LS174T/S cells to LS174T/R cell-derived Exo increased the expression of Nrf2 and P-glycoprotein (P-gp) compared to the nontreated LS174T/S cells (p < .05). In line with these changes, lower intracellular Rhodamin 123 content was detected in Exo-treated cells compared to the nontreated LS174T/S cells. Exo increased migration and clonogenic capacity of LS174T/S cells after incubation with Exo-derived from resistant cells. Of note, inhibition of Nrf2 with a specific blocker, brusatol, blunted these effects. Taken together, Exo-mediated transfer of Nrf2 is involved in the development of oxaliplatin resistance in CC cells by upregulating P-gp.

Oncostatin M and its receptor in women with polycystic ovary syndrome and association with assisted reproductive technology outcomes.

The role of adipokines in ovarian-related disorders such as polycystic ovary syndrome (PCOS) has been reported. However, the involvement of Oncostatin M (OSM), a recently identified adipokine, in ovarian function is unknown. Therefore, we investigated the association of the OSM signaling pathway with ovarian functions and PCOS pathogenesis. This case-control study enrolled 30 PCOS and 30 healthy women who underwent the intracytoplasmic sperm injection procedure. OSM and OSM receptor (OSMR) levels were evaluated in the follicular fluid (FF). Moreover, the expression of insulin receptor substrates (IRS1 and IRS2), OSM, OSMR, suppressor of cytokine signaling 3 (SOCS3), and androgen receptor (AR) genes were analyzed in the isolated cumulus cells (CCs). For the in-vitro experiment, the effect of recombinant OSM on the expression of related genes in isolated CCs was analyzed. Follicular concentrations of OSM and OSMR were significantly lower in PCOS (123.91±48.58 pg/ml and 0.93±0.35 ng/ml, respectively) compared to control women (283.53 ± 96.62 pg/ml and 1.45 ± 0.18 ng/ml, respectively; p < 0.001) and were positively correlated with the oocyte maturation (r = 0.611 and r = 0.611, respectively) and fertilization (r = 0.592 and r = 0.627, respectively) rates in the PCOS group. Furthermore, the SOCS3 expression was upregulated about eight times in PCOS patients compared to the controls (p < 0.05). The treatment of cells with recombinant OSM significantly increased SOCS3, OSMR, IRS-1, and -2 expression and decreased AR expression. The decreased levels of OSM and its receptor in PCOS patients, possibly mediated by SOCS3, could negatively affect oocyte maturation and fertilization rates.

Thymoquinone inhibited vasculogenic capacity and promoted mesenchymal-epithelial transition of human breast cancer stem cells.

BACKGROUND: Vasculogenic mimicry (VM) is characterized by the formation of tubular structure inside the tumor stroma. It has been shown that a small fraction of cancer cells, namely cancer stem cells (CSCs), could stimulate the development of vascular units in the tumor niche, leading to enhanced metastasis to the remote sites. This study aimed to study the inhibitory effect of phytocompound, Thymoquinone (TQ), on human breast MDA-MB-231 cell line via monitoring Wnt/PI3K signaling pathway. METHODS: MDA-MB-231 CSCs were incubated with different concentrations of TQ for 48 h. The viability of CSCs was determined using the MTT assay. The combination of TQ and PI3K and Wnt3a inhibitors was examined in CSCs. By using the Matrigel assay, we measured the tubulogenesis capacity. The percent of CD24- CSCs and Rhodamine 123 efflux capacity was studied using flow cytometry analysis. Protein levels of Akt, p-Akt, Wnt3a, vascular endothelial-cadherin (VE-cadherin), and matrix metalloproteinases-2 and -9 (MMP-2 and -9) were detected by western blotting. RESULTS: TQ decreased the viability of CSCs in a dose-dependent manner. The combination of TQ with PI3K and Wnt3a inhibitors reduced significantly the survival rate compared to the control group (p < 0.05). TQ could blunt the stimulatory effect of vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), fibroblast growth factor (FGF) on CSCs (p < 0.05). The vasculogenic capacity of CSCs was reduced after being-exposed to TQ (p < 0.05). Western blotting revealed the decrease of CSCs metastasis by suppressing MMP-2 and -9. The protein level of VE-cadherin was also diminished in TQ-treated CSCs as compared to the control cell (p < 0.05), indicating inhibition of mesenchymal-endothelial transition (MendT). TQ could suppress Wnt3a and PI3K, which coincided with the reduction of the p-Akt/Akt ratio. TQ had the potential to decrease the number of CD24- CSCs and Rhodamine 123 efflux capacity after 48 h. CONCLUSION: TQ could alter the vasculogenic capacity and mesenchymal-epithelial transition of human breast CSCs in vitro. Thus TQ together with anti-angiogenic therapies may be a novel therapeutic agent in the suppression of VM in breast cancer.

Potential roles and prognostic significance of exosomes in cancer drug resistance.

Drug resistance is a major impediment in cancer therapy which strongly reduces the efficiency of anti-cancer drugs. Exosomes are extracellular vesicles with cup or spherical shape with a size range of 40-150 nm released by eukaryotic cells that contain genetic materials, proteins, and lipids which mediate a specific cell-to-cell communication. The potential roles of exosomes in intrinsic and acquired drug resistance have been reported in several studies. Furthermore, a line of evidence suggested that the content of exosomes released from tumor cells in biological samples may be associated with the clinical outcomes of cancer patients. In this review, we highlighted the recent studies regarding the potential roles of exosomes in tumor initiation, progression, and chemoresistance. This study suggests the possible role of exosomes for drug delivery and their contents in prognosis and resistance to chemotherapy in cancer patients.

Inhibitory effect of melatonin on hypoxia-induced vasculogenic mimicry via suppressing epithelial-mesenchymal transition (EMT) in breast cancer stem cells.

Vasculogenic mimicry (VM) play an important role in breast cancer metastasis and anti- angiogenic drugs resistance. Hypoxia, the epithelial-mesenchymal transition (EMT), and cancer stem cells (CSCs) are known as essential factors for VM formation. Also, melatonin is an amino acid-derived hormone with many anti-tumor effects. Despite the antitumor effects of melatonin, its effect on VM formation in breast cancer has not been considered yet, so we investigated the effect of melatonin on VM formation through EMT process under hypoxia conditions in breast CSCs. The CSCs percentage and VM formation were determined in MCF-7 and MDA-MB-231, respectively. Also, analysis of HIF-1α expression under hypoxia in MDA-MB-231 and MCF-7 cell lines was performed using Western blot. The effect of melatonin on the VM formation, invasion, and migration was also investigated. Moreover, the effect of melatonin on the expression EMT markers was evaluated. CD44+ CD24-phenotype as CSCs marker in MDA-MB-231 cell line, was 80.8%, while it was 11.1% in MCF-7 cell line. HIF-1α expression was up-regulated in the VM-positive breast cancer cell line MDA-MB-231, and consequently, affected the expression of the EMT markers E-cadherin, vimentin, snail, and MMP9. Melatonin had significant effect on EMT and formations of VM in breast CSCs. Melatonin could prevent the formation of VM by affecting the important molecules involved in the formation of VM structures and the EMT. Moreover, our data clearly showed that, melatonin represents molecule with significant anti-cancer activities that may potentially optimize the management of breast cancer through the overcoming drug resistance in anti-angiogenic drugs.

Targeting vasculogenic mimicry by phytochemicals: A potential opportunity for cancer therapy.

Vasculogenic mimicry (VM) is regarded as a process where very aggressive cancer cells generate vascular-like patterns without the presence of endothelial cells. It is considered as the main mark of malignant cancer and has pivotal role in cancer metastasis and progression in various types of cancers. On the other hand, resistance to the antiangiogenesis therapies leads to the cancer recurrence. Therefore, development of novel chemotherapies and their combinations is urgently needed for abolition of VM structures and also for better tumor therapy. Hence, identifying compounds that target VM structures might be superior therapeutic factors for cancers treatment and controlling the recurrence and metastasis. In recent times, naturally occurring compounds, especially phytochemicals have obtained great attention due to their safe properties. Phytochemicals are also capable of targeting VM structure and also their main signaling pathways. Consequently, in this review article, we illustrated key signaling pathways in VM, and the phytochemicals that affect these structures including curcumin, genistein, lycorine, luteolin, columbamine, triptolide, Paris polyphylla, dehydroeffusol, jatrorrhizine hydrochloride, grape seed proanthocyanidins, resveratrol, isoxanthohumol, dehydrocurvularine, galiellalactone, oxacyclododecindione, brucine, honokiol, ginsenoside Rg3, and norcantharidin. The recognition of these phytochemicals and their safety profile may lead to new therapeutic agents' development for VM elimination in different types of tumors.

ß-LAPachone ameliorates doxorubicin-induced cardiotoxicity via regulating autophagy and Nrf2 signalling pathways in mice.

ß-LAPachone (B-LAP) is a naphthoquinone that possesses antioxidant properties. In the present investigation, the protective effect of B-LAP against doxorubicin (DOX)-induced cardiotoxicity was examined in mice. Thirty-five mice were divided into 5 groups: control group, B-LAP (5 mg/kg) group, DOX (15 mg/kg) group, DOX+B-LAP (2.5 mg/kg) group and DOX+B-LAP (5 mg/kg) group. B-LAP was administered orally for 14 days of experimental period. A single dose of DOX (15 mg/kg) was injected intraperitoneally on day 3. Cardiac function, histoarchitecture, indices of oxidative stress and circulating markers of cardiac injury were examined. B-LAP (5 mg/kg) decreased serum levels of lactate dehydrogenase (LDH), creatine kinase MB (CK-MB) and cardiac troponin I (cTnI), and ameliorated cardiac histopathological alterations. In addition to increasing cellular NAD+ /NADH ratio, B-LAP up-regulated the cardiac levels of SIRT1, beclin-1, p-LKB1 and p-AMPK, and reduced the cardiac levels of p-mTOR, interleukin (IL)-1ß, TNF (tumour necrosis factor)-α and caspase-3. B-LAP also elevated the nuclear accumulation of Nrf2 and simultaneously up-regulated the protein levels of haem oxygenase (HO-1) and glutathione S-transferase (GST) in the hearts of DOX mice. While B-LAP reduced malondialdehyde concentrations in heart of DOX-treated mice, it further promoted the activities of cardiac superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT).In accordance with increased cell survival, B-LAP significantly improved the cardiac function of DOX mice. Collectively, these findings underline the protective potential of B-LAP against DOX-induced cardiotoxicity by regulating autophagy and AMPK/Nrf2 signalling pathway in mice.

The effect of oral melatonin on renal ischemia-reperfusion injury in transplant patients: A double-blind, randomized controlled trial.

BACKGROUND: One of the important factors in the occurrence of acute kidney injury (AKI) among renal transplant patients (RTPs) is ischemia reperfusion injury (IRI). The current study aimed at determining the anti-inflammatory and anti-oxidative effects of melatonin on the complications of IRI and the level of Klotho expression in these patients. METHODS: A total of 40 renal transplant candidates were randomly assigned into placebo or melatonin group receiving the same dose of 3 mg/day. In order to measure serum melatonin levels, inflammatory and oxidative stress factors, renal function biomarkers, and Klotho gene/protein expression, venous blood samples were taken from patients over two different time points, i e, 24 h before the transplantation and at discharge from hospital. RESULTS: Melatonin was associated with improvement in renal transplantation, since the serum level of neutrophil gelatinase-associated lipocalin, as a renal functional marker, significantly decreased (P < .001). The effect of melatonin as a suppressor of inflammation and oxidative stress was also evident in the melatonin group due to a significant reduction in the serum levels of MDA, CP, 8-OHdG, and TNF-α markers (P < .001). CONCLUSIONS: Reduction in serum levels of renal function and oxidative stress/inflammatory markers in the melatonin group indicates that melatonin can inhibit IRI outcomes in RTPs through its anti-oxidant and anti-inflammatory properties. However, these properties do not appear as a result of influence on the level of Klotho gene/protein expression.