NIH 3T3 Mouse Embryonic Fibroblast Research Articles

DNMT1 driven by mouse amniotic fluid mesenchymal stem cell exosomes improved corneal cryoinjury via inducing microRNA-33 promoter DNA hypermethylation modification in corneal epithelium cells.

Severe corneal cryoinjury can cause permanent corneal swelling and bullous keratopathy, one of the main reason for loss of sight. Mouse amniotic fluid mesenchymal stem cells (mAF-MSCs) can repair corneal damage caused by freezing; however, whether the exosomes derived from mAF-MSCs have the same repair effect is unknown. In this study, the mAF-MSC-exosomes were transplanted into the eyeballs of corneal cryoinjured mice. Histopathological examination showed that the mAF-MSC-exosomes improved the corneal structure and status of corneal epithelial cells in corneal cryoinjured mice. RRBS-sequencing showed that compared with the control group, four genes (Rpl13-ps6, miR-33, Hymai, and Plagl1), underwent DNA hypermethylation modification after mAF-MSC-exosomes treatment. The result of FISH indicated that miR-33-3p hybridization signals were enhanced in corneal epithelial cells from mice treated with mAF-MSC-exosomes. Semi-quantitative PCR and western blotting indicated that mAF-MSC-exosomes contained high levels of DNMT1 mRNA and protein. Additionally, luciferase report assays indicated that miR-33-3p overexpression in NIH-3T3 mouse embryonic fibroblast cells inhibited the activity of luciferase carrying a sequence from the 3' untranslated region of Bcl6. Moreover, BCL6 mRNA and protein levels in corneal tissues from mice treated with mAF-MSC-exosomes were higher than those in the control group. Therefore, our results suggested that mAF-MSC-exosomes could repair corneal cryoinjury by releasing DNMT1, which induced hypermethylation of the miR-33 promoter in corneal epithelial cells. Consequent downregulated miR-33 transcription upregulated Bcl6 expression, ultimately achieving the repair of corneal cryoinjury in mice.

Synthesis, Cytotoxic Activity Evaluation and Molecular Docking Studies of Some Benzimidazole Derivatives

In this study, the synthesis of 2-(2-acetyl-1H-benzimidazol-1-yl)-1-arylethanone (3a-3d) and 1-methyl-3-phenyl-benzo[4,5]imidazo[1,2-a]pyrazine derivatives (4a-4d) and to investigate their cytotoxic activity were aimed. APCI, IR, 1HNMR, and 13CNMR spectra were utilized to determine the structure of the synthesized compounds. The cytotoxic activity of selected compounds were detected in A549 (human lung carcinoma) and NIH3T3 (mouse embryonic fibroblasts) cell lines. Compounds 4c and 4d were found to be selectively cytotoxic against A549 and NIH3T3 cell lines. Molecular docking studies were performed using the data retrieved from the Protein Data Bank server (PDBID: 4QTX).

A Series of Non-Oxido VIV Complexes of Dibasic ONS Donor Ligands: Solution Stability, Chemical Transformations, Protein Interactions, and Antiproliferative Activity.

A series of mononuclear non-oxido vanadium(IV) complexes, [VIV(L1-4)2] (1-4), featuring tridentate bi-negative ONS chelating S-alkyl/aryl-substituted dithiocarbazate ligands H2L1-4, are reported. All the synthesized non-oxido VIV compounds are characterized by elemental analysis, spectroscopy (IR, UV-vis, and EPR), ESI-MS, as well as electrochemical techniques (cyclic voltammetry). Single-crystal X-ray diffraction studies of 1-3 reveal that the mononuclear non-oxido VIV complexes show distorted octahedral (1 and 2) or trigonal prismatic (3) arrangement around the non-oxido VIV center. EPR and DFT data indicate the coexistence of mer and fac isomers in solution, and ESI-MS results suggest a partial oxidation of [VIV(L1-4)2] to [VV(L1-4)2]+ and [VVO2(L1-4)]-; therefore, all these three complexes are plausible active species. Complexes 1-4 interact with bovine serum albumin (BSA) with a moderate binding affinity, and docking calculations reveal non-covalent interactions with different regions of BSA, particularly with Tyr, Lys, Arg, and Thr residues. In vitro cytotoxic activity of all complexes is assayed against the HT-29 (colon cancer) and HeLa (cervical cancer) cells and compared with the NIH-3T3 (mouse embryonic fibroblast) normal cell line by MTT assay and DAPI staining. The results suggest that complexes 1-4 are cytotoxic in nature and induce cell death in the cancer cell lines by apoptosis and that a mixture of VIV, VV, and VVO2 species could be responsible for the biological activity.

Synthesis and Antitumor Evaluation of Biotin-SN38-Valproic Acid Conjugates.

Despite the strong anticancer activity of SN38 (7-ethyl-10-hydroxy-camptothecin), the severe side effects and loss of anticancer activity caused by the lack of selectivity to cancer cells and hydrolysis of ring E prevent its clinical application. To address the issue, herein a multifunctional SN38 derivative (compound 9) containing biotin (tumor-targeting group) and valproic acid (histone deacetylase inhibitor, HDACi) was synthesized via click chemistry and evaluated using MTT assay. The in vitro cytotoxicity study showed that compound 9 exhibited superior cytotoxicity than irinotecan against human cervical cancer HeLa cells, albeit it was inferior to SN38. More significantly, compound 9 significantly reduced toxicity in mouse embryonic fibroblast NIH3T3 cells, indicating that compound 9 had the capacity to enhance tumor targeting due to its cell selectivity. Further studies demonstrated that, compared with irinotecan, compound 9 induced similar apoptosis of cancer cells. Consequently, compound 9 can not only improve its tumor-targeting ability mediated by biotin but also exert potent anticancer activity through the effect of SN38 and valproic acid, indicating that the design concept is an effective strategy for the structural modification of SN38.

UV-Accelerated Synthesis of Gold Nanoparticle-Pluronic Nanocomposites for X-ray Computed Tomography Contrast Enhancement.

Eco-friendly chemical methods using FDA-approved Pluronic F127 (PLU) block copolymer have garnered much attention for simultaneously forming and stabilizing Au nanoparticles (AuNPs). Given the remarkable properties of AuNPs for usage in various fields, especially in biomedicine, we performed a systematic study to synthesize AuNP-PLU nanocomposites under optimized conditions using UV irradiation for accelerating the reaction. The use of UV irradiation at 254 nm resulted in several advantages over the control method conducted under ambient light (control). The AuNP-PLU-UV nanocomposite was produced six times faster, lasting 10 min, and exhibited lower size dispersion than the control. A set of experimental techniques was applied to determine the structure and morphology of the produced nanocomposites as affected by the UV irradiation. The MTT assay was conducted to estimate IC50 values of AuNP-PLU-UV in NIH 3T3 mouse embryonic fibroblasts, and the results suggest that the sample is more compatible with cells than control samples. Afterward, in vivo maternal and fetal toxicity assays were performed in rats to evaluate the effect of AuNP-PLU-UV formulation during pregnancy. Under the tested conditions, the treatment was found to be safe for the mother and fetus. As a proof of concept or application, the synthesized Au:PLU were tested as contrast agents with an X-ray computed tomography scan (X-ray CT).

Developing a Reliable Holographic Flow Cyto-Tomography Apparatus by Optimizing the Experimental Layout and Computational Processing.

Digital Holographic Tomography (DHT) has recently been established as a means of retrieving the 3D refractive index mapping of single cells. To make DHT a viable system, it is necessary to develop a reliable and robust holographic apparatus in order that such technology can be utilized outside of specialized optics laboratories and operated in the in-flow modality. In this paper, we propose a quasi-common-path lateral-shearing holographic optical set-up to be used, for the first time, for DHT in a flow-cytometer modality. The proposed solution is able to withstand environmental vibrations that can severely affect the interference process. Furthermore, we have scaled down the system while ensuring that a full 360° rotation of the cells occurs in the field-of-view, in order to retrieve 3D phase-contrast tomograms of single cells flowing along a microfluidic channel. This was achieved by setting the camera sensor at 45° with respect to the microfluidic direction. Additional optimizations were made to the computational elements to ensure the reliable retrieval of 3D refractive index distributions by demonstrating an effective method of tomographic reconstruction, based on high-order total variation. The results were first demonstrated using realistic 3D numerical phantom cells to assess the performance of the proposed high-order total variation method in comparison with the gold-standard algorithm for tomographic reconstructions: namely, filtered back projection. Then, the proposed DHT system and the processing pipeline were experimentally validated for monocytes and mouse embryonic fibroblast NIH-3T3 cells lines. Moreover, the repeatability of these tomographic measurements was also investigated by recording the same cell multiple times and quantifying the ability to provide reliable and comparable tomographic reconstructions, as confirmed by a correlation coefficient greater than 95%. The reported results represent various steps forward in several key aspects of in-flow DHT, thus paving the way for its use in real-world applications.

ATRT-24. CDK7 Inhibition in AT/RT

Atypical teratoid/rhabdoid tumors (AT/RT) are CNS tumors with a 5-year survival of ~35%. AT/RT is characterized by loss-of-function mutations in the SMARCB1 component of the SWI/SNF (SWItch/Sucrose Non-Fermentable) complex. Based on preliminary CRISPR-Cas9 gene essentiality screen results identifying AT/RT vulnerabilities, we hypothesized that interaction between CDK7 and the SWI/SNF complex via SMARCB1 provides a potential target to improve clinical survival of patients. CDK7 expression was identified by microarray in AT/RT, medulloblastoma, glioblastoma and normal brain. Established cell lines (BT12, BT16, CHLA06), patient derived lines (MAF-737, MAF-1298, MAF-1337), normal human astrocytes (NHA) and NIH3T3 mouse embryonic fibroblast cells were utilized for in vitro response to CDK7 inhibition. Murine cerebellar xenografts of MAF-737 were utilized to evaluate genetic and pharmacologic response to CDK7 inhibition. The NCI Approved Oncology Drugs (AOD-9) Panel was evaluated with an IC25 dose of CDK7 inhibitor THZ2 to identify potential synergistic combinations. CDK7 is up-regulated in AT/RT compared to other brain tumors or normal brain. In vitro, AT/RT cells are highly susceptible to CDK7 pharmacologic inhibition with nM IC50 levels. AT/RT cells with shRNA against CDK7 implanted in vivo show significantly reduced growth. Evaluation of in vivo tumors treated with THZ2 demonstrate decreased Ki-67 and reduced pRBP1 demonstrating effective inhibition of the target as well as a decrease in cell proliferation. Combination therapy of THZ2 with the AOD-9 Panel found significant synergy with antimetabolite therapies, specifically pemetrexed, pralatrexate, and methotrexate. There was no synergy with other standard chemotherapy. Our findings demonstrate that CDK7 is highly expressed in AT/RT and necessary for proliferation of AT/RT cells, suggesting it as a potential therapeutic target. Antimetabolites, which are currently used in several AT/RT protocols, synergized with CDK7 inhibition offers a potential future combination therapy for patients.

Recombinant mussel protein Pvfp5β enhances cell adhesion of poly(vinyl alcohol)/k-carrageenan hydrogel scaffolds

Polymeric hydrogels are increasingly considered as scaffolds for tissue engineering due to their extraordinary resemblance with the extracellular matrix (ECM) of many tissues. As cell adhesion is a key factor in regulating important cell functions, hydrogel scaffolds are often functionalized or loaded with a variety of bioactive molecules that can promote adhesion. Interesting biomimetic approaches exploit the properties of mussel-inspired recombinant adhesive proteins. In this work, we prepared hydrogel scaffolds with a 50%w mixture of k-carrageenan (kC) and polyvinyl alcohol (PVA), by a two-step physical gelation process, and we coated them with Perna viridis foot protein-5β (Pvfp5β). The mechanical and morphological properties of hydrogels were investigated both after conditioning with typical cell culture media and also after coating with the Pvfp5β. The protein resulted strongly adsorbed onto the surface of the hydrogel and also able to penetrate in its interiors to a certain depth, mainly interacting with the kC component of the scaffold as resulted from the confocal analysis. Mouse embryonic fibroblasts NIH-3T3 were seeded on top of the hydrogels and cultured up to two weeks. The role of Pvfp5β in promoting cell adhesion, spreading and colonization of the scaffold was demonstrated.

The in vitro Evaluation of Protective Effect of Rutin on Acrylamide Induced Oxidative Aging in NIH3T3 Cells

Background: Aging is one of the important factor in the development of age-related diseases. Acrylamide (ACR) can be produced during rich-carbohydrate foods preparing at high temperature. Recently, studies showed that ACR can induce cellular aging. Furthermore, Rutin is a natural flavonoid, which possesses a potent antioxidant activity. Objective: In this study, ptotective effect of Rutin (a potent antioxidant) on oxidative-induced aging after ACR exposure was evaluated. Materials and Methods: NIH3T3 mouse embryonic fibroblast cells were used for this study, which treated by different concentrations of ACR and H2O2. Oxidative stress was assessed by measuring of malondialdehyde and glutathion concentrations. To evaluate of the aging process, β-galactosidase activity was evaluated by enzyme staining and β-galactosidase activity assay kit. MTT assay was used to evaluate the viability of the cells. Results: Exposure of cells to ACR and H2O2 significantly decreased the cell viability and using of rutin significantly improved cell viability in cells treated by ACR (P<0.05). Rutin increased glutathion level in cells treated by ACR and H2O2 (P<0.05). Also, the rate of lipid peroxidation was significantly lower in the group treated with rutin and H2O2 than in the H2O2 group (P<0.05).The β -galactosidase activity was increased in H2O2 and ACR groups. The using of rutin with acrylamide significantly reduced the activity of β-galactosidase enzyme compared to acrylamide group (P<0.05). Conclusion: Based on this study, oxidative stress may be presented as an important mechanism in ACR induced-aging in NIH3T3 cells. On the other hand, Rutin, as a potent antioxidant, decresead aging by inhibition of oxidative damage.

A New System of Sustainable Silico-Aluminous and Silicate Materials for Cultivation Purpose within Sustainable Buildings: Chemical-Physical, Antibacterial and Cytotoxicity Properties

In this study, we compared the chemical-physical, antibacterial, and cytotoxicity properties of silico-aluminous and silicate materials for outdoor (green roof, planted walls) and indoor (urban farms, indoor microgreen gardens) cultivation purpose in a context of sustainable construction. Glasses and lightweight aggregates were tailored starting from waste, by-product, and post-consumer and bioproducts (packaging glass cullet, cattle bone flour ash, vegetable biomass ash, spent coffee ground, degreased from biomass of prepupae of Black Soldier Flies) mixed together with a national ferruginous red clay, quarry scrap pumice and, if necessary, with K2CO3 of reagent grade. The first type of material was obtained by melting at 1200 °C and the second one by powder sintering at 1000 °C. All specimens, subjected to antibacterial test, showed both low zone of inhibitions towards two Gram-negative and two Gram-positive bacterial strains. A cytotoxicity test on mouse embryonic fibroblast NIH-3T3 cell line directly exposed to the investigated materials was performed at three different exposure times (1 h, 3 h, and 6 h). Data acquired highlighted that the materials positively affected redox mitochondrial activity of the fibroblast cells. The concentrations of leachate heavy metals detected on selected materials in water at room temperature after 24 h were lower than the European law limit and an interesting release of P, K, and N nutrients was noted for those formulations designed for agronomic purposes. pH, falling on average within the 6.5–7.5 range, is optimal for most crops, and the specific conductivity <2 dS/m indicates no depression danger for crops. Both bulk density <1200 kg/m3 and porosity over 50% seem to ensure good performance of lightening, drainage, water reservation, and oxygenation of the roots.

In vitro studies on the cytotoxicity, elastase, and tyrosinase inhibitory activities of tomato (Solanum lycopersicum Mill.) extract.

ABSTRACTTomatoes (Solanum lycopersicum Mill.), a common vegetable in Indonesia, contain high levels of lycopene, which is good for the body. This research further investigates the activity of polar and nonpolar fractions of tomatoes as elastase and tyrosinase inhibitory, and cytotoxic agents. The extraction procedure used is maceration, fractionation through liquid-liquid fractionation, purification of phytochemical substances is achieved through the application of thin layer chromatography. Elastase and tyrosinase inhibitory activity was analyzed using spectrophotometry and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cytotoxic assay. The result showed that the extract yield was 0.004%. The percentage of polar fraction from the extract was 2.58%, while the nonpolar fraction was 0.69%. The elastase inhibitory activity of polar and nonpolar fractions of tomato extract is 87.21% ± 7.57% and 73.12% ± 7.44%, respectively, The elastase inhibitory activity of polar and nonpolar fractions of tomato extract is 87.21% ± 7.57% and 73.12% ± 7.44%, respectively. The fractions had higher the anti-elastase activity than the positive control quercetin (65.97% ± 3.00%). The tyrosinase inhibitory activity of polar and nonpolar fractions of tomato extract is 23.71% ± 7.91% and 41.16% ± 5.41% (kojic acid as standard is 65.07% ± 0.86%), respectively. The IC50 of the cytotoxic assay to NIH 3T3 mouse embryonic fibroblast cells of the polar and nonpolar fraction of tomato extract is 1820.90 μg/mL and 1643.86 μg/mL, respectively.

Antibacterial Properties and Cytotoxicity of 100% Waste Derived Alkali Activated Materials: Slags and Stone Wool-Based Binders

In this study we compare the leaching behavior and the antibacterial and cytotoxic properties of 100% slag or stone wool derived alkali activated materials. The antibacterial activity was measured as the inhibiting capacity against two Gram-negative bacterial strains, Escherichia coli and Pseudomonas aeruginosa and one Gram-positive bacterial strain: Enterococcus faecalis. The cytotoxicity properties were tested on mouse embryonic fibroblast NIH-3T3 cell-line. It was proved that the high quality of the 3D aluminosilicate network of the consolidated materials obtained from powders of CaO or MgO-rich slags or stone wool, opportunely activated with NaO and/or Na-silicate, was capable of stabilizing heavy metal cations. The concentrations of leachate heavy cations were lower than the European law limit when tested in water. The effect of additives in the composites, basal fibers or nanocellulose, did not reduce the chemical stability and slightly influenced the compressive strength. Weight loss in water increased by 20% with basalt fibers addition, while it remained almost constant when nanocellulose was added. All the consolidated materials, cement-like in appearance, exhibited limited antibacterial properties (viability from 50 to 80% depending on the bacterial colony and the amount of sample) and absence of cytotoxicity, envisaging good acceptance from part of the final consumer and zero ecological impact. CaO-rich formulations can replace ordinary Portland cement (showing bacterial viability at 100%) with a certain capability for preventing the reproduction of the E. coli and S. aureus bacteria with health and environmental protection results.

Short antimicrobial peptidomimetic SAMP-5 effective against multidrug-resistant gram-negative bacteria

SAMP-5 is a short histidine-derived antimicrobial peptidomimetic with pendant dialkylated tail. In this study, we evaluated the potential of SAMP-5 as an antimicrobial agent to combat multidrug-resistant gram-negative bacteria. SAMP-5 showed potent antimicrobial activity (minimum inhibitory concentration 16-64 μg/ml) comparable to melittin against multidrug-resistant Escherichia coli (MDREC) and multidrug-resistant (MDRPA). SAMP-5 displayed no cytotoxicity against three mammalian cells such as mouse macrophage RAW264.7, mouse embryonic fibroblast NIH-3T3, and human bone marrow SH-SY5Y cells at the concentration of 128 μg/ml. SAMP-5 showed resistance to proteolytic degradation with pepsin, trypsin, α-chymotrypsin, and proteinase K. Importantly, unlike ciprofloxacin, no antibiotic resistance against SAMP-5 arose for Pseudomonas aeruginosa during 7 days of serial passage at 0.5 × MIC. Moreover, SAMP-5 showed synergy or additive effects against MDRPA and MDREC, when it combined with chloramphenicol, ciprofloxacin, and oxacillin. Collectively, our results suggested that SAMP-5 is a promising alternative and adjuvant to treat infections caused by multidrug-resistant gram-negative bacteria.

MicroRNA-26 regulates the expression of CTGF after exposure to ionizing radiation.

Radiation-induced fibrosis (RIF) is a serious complication that occurs after irradiation and which is caused by the deposition of extracellular matrix (ECM) proteins such as collagen. However, the underlying mechanisms, including the expression of the cytokines, that promote the RIF process, are not yet fully understood. MicroRNAs (miRNAs) have recently been suggested to act as post-transcriptional repressors for many genes; however, their role in the process of RIF remains to be elucidated. Our previous study showed that ionizing radiation increased the type I collagen expression through the activation of transforming growth factor (TGF)-β, while miR-29 repressed this increase. This study aimed to investigate the mechanisms by which the expression of connective tissue growth factor (CTGF), a downstream mediator of TGF-β, is controlled by miRNAs post-transcriptionally after exposure to ionizing radiation. The expression of CTGF in NIH-3T3 cells and mouse embryonic fibroblasts was increased by ionizing radiation. However, this increase was suppressed with a specific inhibitor of TGF-β receptor. Among the predictable miRNAs that target the CTGF gene, the expression of miR-26a was downregulated after exposure to ionizing radiation and this regulation was negatively mediated by TGF-β signaling. miR-26a negatively regulated the CTGF expression at the post-transcriptional level; however, ionizing radiation suppressed this negative regulation. In addition, the overexpression of miR-26a inhibited the expression of CTGF and type I collagen after irradiation. In conclusion, miR-26a modulates the expression of CTGF via TGF-β signaling in irradiated fibroblasts. The results suggest the potential application of miR-26a in the treatment of RIF.

Studies of anticancer activity in vivo and in vitro behaviors of liposomes encapsulated iridium(III) complex.

Iridium(III) complexes have gained great attention in cancer treatment in recent years. In this paper, we designed and synthesized a new iridium(III) complex [Ir(piq)2(DQTT)](PF6) Ir1 (piq = 1-phenylisoquinoline, DQTT = 12-(1,4-dihydroquinoxalin-6-yl)-4,5,9,14-tetraazabenzo[b]triphenylene). The Ir1-loaded PEGylated liposomes (Lipo-Ir1) were prepared using the ethanol injection method. The anticancer activity of the complex and Lipo-Ir1 against SGC-7901 (human gastric adenocarcinoma), A549 (human lung carcinoma), HeLa (human cervical carcinoma), HepG2 (human hepatocellular carcinoma), BEL-7402 (human hepatocellular carcinoma), and normal NIH3T3 (mouse embryonic fibroblasts) was tested by the MTT method. The complex Ir1 shows moderate or low cytotoxicity against the selected cancer cells, whereas the Lipo-Ir1 exhibits high anticancer activity toward the same cancer cells. The apoptosis induced by Lipo-Ir1 was assayed by flow cytometry and Lipo-Ir1 induced apoptosis through increasing intracellular reactive-oxygen species levels, decreasing mitochondrial membrane potential, further promoting cytochrome c release and causing the increase of level of intracellular Ca2+. Western blot was used to detect the changes in Bcl-2 family protein and PI3K/AKT pathway proteins. The cloning experiments demonstrated that the Lipo-Ir1 can effectively inhibit cell proliferation. In vivo experiments, Lipo-Ir1 inhibited tumor growth in xenograft nude mice, and the percentage of tumor growth inhibition in vivo was 75.70%. Overall, the liposomes Lipo-Ir1 exhibits higher anticancer activity than Ir1 under the same conditions. These results indicated that Lipo-Ir1 may be a valuable resource for cancer therapy.

Exploring anticancer efficiency of mitochondria-targeted cyclometalated iridium(III) complexes

We prepared and characterized new iridium(III) complexes: [Ir(NC)2(MPPIP)](PF6) (N-C = 2-phenylpyridine 1; benzo[h]quinolone 2; 1-phenylisoquinolone, 3, MPPIP = 2-(4-(4′-methylpiperazin-yl)phenyl)-1H-imidazo[4,5-f][1,10]phenanthroline). MTT (MTT = 3-(4,5-dimethylthiazole-2-yl)-2,5-biphenyl tetrazolium bromide) method was used to assay anticancer activities of the complexes 1-3 toward SGC-7901, HeLa, A549, BEL-7402, mouse embryonic fibroblast NIH3T3 cell lines. Complexes 1, 2, 3 are sensitive to A549 cells and display a relatively low IC50 value of 5.4 ± 0.3, 4.2 ± 0.03 and 3.8 ± 0.2 μM, respectively. The apoptotic efficiency was investigated and the number of apoptotic cells induced by 1, 2 and 3 is 9.92%, 11.30% and 16.00%. The complexes are able to increase intracellular ROS content and lessen the mitochondrial membrane potential. Besides, anti-tumor activity in vivo reveals that complex 3 exhibits moderate effect on inhibiting the tumor growth, and complex 3 has no influence on liver, brain, kidney, lung and heart.

Chitosan-Based Bioactive Glass Gauze: Microstructural Properties, In Vitro Bioactivity, and Biological Tests

Passive commercial gauzes were turned into interactive wound dressings by impregnating them with a chitosan suspension. To further improve healing, and cell adhesion and proliferation, chitosan/bioactive glass wound dressings were produced with the addition of (i) 45S5, (ii) a Sr- and Mg-containing bioactive glass, and (iii) a Zn-containing bioactive glass to the chitosan suspension. SEM and FTIR analyses evidenced positive results in terms of incorporation of bioactive glass particles. Bioactivity was investigated by soaking chitosan-based bioactive glass wound dressings in simulated body fluid (SBF). Cell viability, proliferation, and morphology were investigated using NIH 3T3 (mouse embryonic fibroblast) cells by neutral red (NR) uptake and MTT assays. Furthermore, the wound-healing rate was evaluated by means of the scratch test, using NIH 3T3. The results showed that bioactive glass particles enhance cell adhesion and proliferation, and wound healing compared to pure chitosan. Therefore, chitosan-based bioactive glass wound dressings combine the properties of the organic matrix with the specific biological characteristics of bioactive glasses to achieve chitosan composites suitable for healing devices.

The Effect of Phase Transition Temperature on Therapeutic Efficacy of Liposomal Bortezomib.

Here, three liposomal formulations of DPPC/DPPG/Chol/DSPE-mPEG2000 (F1), DPPC/DPPG/Chol (F2) and HSPC/DPPG/Chol/DSPE-mPEG2000 (F3) encapsulating BTZ were prepared and characterized in terms of their size, surface charge, drug loading, and release profile. Mannitol was used as a trapping agent to entrap the BTZ inside the liposomal core. The cytotoxicity and anti-tumor activity of formulations were investigated in vitro and in vivo in mice bearing tumor. Bortezomib (BTZ) is an FDA approved proteasome inhibitor for the treatment of mantle cell lymphoma and multiple myeloma. The low solubility of BTZ has been responsible for the several side effects and low therapeutic efficacy of the drug. Encapsulating BTZ in a nano drug delivery system; helps overcome such issues. Among NDDSs, liposomes are promising diagnostic and therapeutic delivery vehicles in cancer treatment. Evaluating anti-tumor activity of bortezomib liposomal formulations. Data prompted us to design and develop three different liposomal formulations of BTZ based on Tm parameter, which determines liposomal stiffness. DPPC (Tm 41°C) and HSPC (Tm 55°C) lipids were chosen as variables associated with liposome rigidity. In vitro cytotoxicity assay was then carried out for the three designed liposomal formulations on C26 and B16F0, which are the colon and melanoma cancer mouse-cell lines, respectively. NIH 3T3 mouse embryonic fibroblast cell line was also used as a normal cell line. The therapeutic efficacy of these formulations was further assessed in mice tumor models. MBTZ were successfully encapsulated into all the three liposomal formulations with a high entrapment efficacy of 60, 64, and 84% for F1, F2, and F3, respectively. The findings showed that liposomes mean particle diameter ranged from 103.4 to 146.8nm. In vitro cytotoxicity studies showed that liposomal-BTZ formulations had higher IC50 value in comparison to free BTZ. F2-liposomes with DPPC, having lower Tm of 41°C, showed much higher anti-tumor efficacy in mice models of C26 and B16F0 tumors compared to F3-HSPC liposomes with a Tm of 55°C. F2 formulation also enhanced mice survival compared with untreated groups, either in BALB/c or in C57BL/6 mice. Our findings indicated that F2-DPPC-liposomal formulations prepared with Tm close to body temperature seem to be effective in reducing the side effects and increasing the therapeutic efficacy of BTZ and merits further investigation.

Effects of Microstripe Geometry on Guided Cell Migration

Cell migration on material surfaces is a fundament issue in the fields of biomaterials, cell biology, tissue engineering, regenerative medicine, etc. Herein, we aim to guide cell migration by flat microstripes with significant contrast of cell adhesion and varied geometric features of the adhesive stripes. To this end, we designed and fabricated cell-adhesive arginine-glycine-aspartate (RGD) microstripes on the nonfouling poly(ethylene glycol) (PEG) background and examined the microstripe-guided adhesion and migration of a few cell types. The migration of cell clusters adhering on the RGD regions was found to be significantly affected by the widths and arc radiuses of the guided microstripes. The cells migrated fastest on the straight microstripes with width of about 20 μm, which we defined as single file confined migration (SFCM). We also checked the possible left-right asymmetric bias of cell migration guided by combinatory microstripes with alternative wavy and quasi-straight stripes under a given width, and found that the velocity of CCW (counter clockwise) migration was higher than that of CW (clockwise) migration for primary rat mesenchymal stem cells (rMSCs), whereas no left-right asymmetric bias was observed for NIH3T3 (mouse embryonic fibroblast cell line) and Hela (human cervix epithelial carcinoma cell line) cells. Comparison of migration of cells on the nanotopological stripe and smooth surfaces further confirmed the importance of cell orientation coherence for guided cell migration and strengthened the superiority of SFCM.

Arthrospira platensis transglutaminase derived antioxidant peptide-packed electrospun chitosan/ poly (vinyl alcohol) nanofibrous mat accelerates wound healing, in vitro, via inducing mouse embryonic fibroblast proliferation

In this present study, we have carried out the antioxidant function of transglutaminase (TG) identified from Arthrospira platensis (Ap) transcriptome. The antioxidant peptide ML11 (MLRSIGIPARL) has been predicted from the transglutaminase core domain and the peptide's free radical scavenging potential was evaluated and it shows that it functions on a dose dependent manner. The ML11 peptide cell toxicity was analysed in the human blood leucocytes which resulted no cytotoxic activity in any of the cell population. Moreover, the nanofibre mat encapsulated with antioxidant peptide ML11 was prepared by electrospinning technique. The antioxidant peptide ML11 encapsulated mat showed increase in fibre diameter compared to the chitosan polyvinyl alcohol blended mat. The change in the crystalline behaviour of both chitosan and polyvinyl alcohol polymer to the amorphous nature was determined by X-ray diffraction at the broad band between 20 and 30° (2θ°). FTIR revealed the functional groups which present in the polymer as well as the interaction between their components of chitosan (CS) and polyvinyl alcohol (PVA). The fibre retains the antioxidant activity due to the peptide encapsulated by scavenging the intracellular ROS that was confirmed by flowcytometry and fluorescence microscopy. The ML11 peptide encapsulated mat showed no cytotoxicity in the NIH-3T3 mouse embryonic fibroblast cells. Also, ML11 peptide encapsulated fibre showed potential wound healing activity in NIH-3T3 cells. Taken altogether, the study indicates that the wound healing potential of the ML11 peptide encapsulated nano fibre mat may be used as biopharmaceutical drug.