Fabricating niosomal-PEG nanoparticles co-loaded with metformin and silibinin for effective treatment of human lung cancer cells

BackgroundDespite current therapies, lung cancer remains a global issue and requires the creation of novel treatment methods. Recent research has shown that biguanides such as metformin (MET) and silibinin (SIL) have a potential anticancer effect. As a consequence, the effectiveness of MET and SIL in combination against lung cancer cells was investigated in this study to develop an effective and novel treatment method.MethodsNiosomal nanoparticles were synthesized via the thin-film hydration method, and field emission scanning electron microscopy (FE-SEM), Fourier transform infrared (FTIR), atomic force microscopy (AFM), and dynamic light scattering (DLS) techniques were used to evaluate their physico-chemical characteristics. The cytotoxic effects of free and drug-loaded nanoparticles (NPs), as well as their combination, on A549 cells were assessed using the MTT assay. An apoptosis test was used while under the influence of medication to identify the molecular mechanisms behind programmed cell death. With the use of a cell cycle test, it was determined whether pharmaceutical effects caused the cell cycle to stop progressing. Additionally, the qRT-PCR technique was used to evaluate the levels of hTERT, BAX, and BCL-2 gene expression after 48-h medication treatment.ResultsIn the cytotoxicity assay, the growth of A549 lung cancer cells was inhibited by both MET and SIL. Compared to the individual therapies, the combination of MET and SIL dramatically and synergistically decreased the IC50 values of MET and SIL in lung cancer cells. Furthermore, the combination of MET and SIL produced lower IC50 values and a better anti-proliferative effect on A549 lung cancer cells. Real-time PCR results showed that the expression levels of hTERT and BCL-2 were significantly reduced in lung cancer cell lines treated with MET and SIL compared to single treatments (p< 0.001).ConclusionIt is anticipated that the use of nano-niosomal-formed MET and SIL would improve lung cancer treatment outcomes and improve the therapeutic efficiency of lung cancer cells

Identification of over producer strain of endo-ß-1,4-glucanase in Aspergillus Species: Characterization of crude carboxymethyl cellulase

Cellulases are a group of hydrolytic enzymes capable of degrading cellulose to smaller sugar components like glucose units. These enzymes are produced by fungi and bacteria. The aim of this research was to identify a Aspergillus species with over production of endo-β-1,4-glucanase. Properties of endo-β-1,4-glucanase/carboxymethylcellulase (CMCase) from a culture filtrate of the Aspergillus sp. was also studied. Aspergillus sp. (R4) was selected as over producer of endo-β-1,4-glucanase among 13 different species. SDS-PAGE activity staining with 1% Congo Red solution revealed three protein bands showing cellulolytic activity. The molecular weights of these proteins were estimated to be approximately 18.5, 23 and 28 kD. Also, conservative region of endo-β-1,4-glucanase coding gene was studied by polymerase chain reaction (PCR). Amplified fragments with 1204 bp and 399 bp were confirmed by restriction pattern with HindII and PstI enzymes. Key Words: Aspergillus sp., Endo-β-1, 4-glucanase, CMCase, SDS-PAGE, PCR. African Journal of Biotechnology Vol.4(1) 2005: 26-3

In Vitro Efficacy of Curcumin-Loaded Amine-Functionalized Mesoporous Silica Nanoparticles against MCF-7 Breast Cancer Cells

Purpose: Mesoporous silica nanoparticles (MSNs) have drawn substantial interest as drug nanocarriers for breast cancer therapy. Nevertheless, because of the hydrophilic surfaces, the loading of well-known hydrophobic polyphenol anticancer agent curcumin (Curc) into MSNs is usually very low. Methods: For this purpose, Curc molecules were loaded into amine-functionalized MSNs (MSNs-NH2 -Curc) and characterized using thermal gravimetric analysis (TGA), Fourier-transform infrared (FTIR), field emission scanning electron microscope (FE-SEM), transmission electron microscope (TEM), Brunauer-Emmett-Teller (BET). MTT assay and confocal microscopy, respectively, were used to determine the cytotoxicity and cellular uptake of the MSNs-NH2 - Curc in the MCF-7 breast cancer cells. Besides, the expression levels of apoptotic genes were evaluated via quantitative polymerase chain reaction (qPCR) and western blot. Results: It was revealed that MSNs-NH2 possessed high values of drug loading efficiency and exhibited slow and sustained drug release compared to bare MSNs. According to the MTT findings, while the MSNs-NH2 -Curc were nontoxic to the human non-tumorigenic MCF-10A cells at low concentrations, it could considerably decrease the viability of MCF-7 breast cancer cells compared to the free Curc in all concentrations after 24, 48 and 72 hours exposure times. A cellular uptake study using confocal fluorescence microscopy confirmed the higher cytotoxicity of MSNs-NH2 -Curc in MCF-7 cells. Further, it was found that the MSNs-NH2 -Curc could drastically affect the mRNA and protein levels of Bax, Bcl-2, caspase 3, caspase 9, and hTERT relative to the free Curc treatment. Conclusion: Taken together, these preliminary results suggest the amine-functionalized MSNs-based drug delivery platform as a promising alternative approach for Curc loading and safe breast cancer treatment

An update of the recombinant protein expression systems of Cyanovirin-N and challenges of preclinical development

Introduction: Human immunodeficiency virus (HIV) is a debilitating challenge and concern worldwide. Accessibility to highly active antiretroviral drugs is little or none for developing countries. Production of cost-effective microbicides to prevent the infection with HIV is a requirement. Cyanovirin-N (CVN) is known as a promising cyanobacterial lectin, capable of inhibiting the HIV cell entry in a highly specific manner. Methods: This review article presents an overview of attempts conducted on different expression systems for the recombinant production of CVN. We have also assessed the potential of the final recombinant product, as an effective anti-HIV microbicide, comparing prokaryotic and eukaryotic expression systems. Results: Artificial production of CVN is a challenging task because the desirable anti-HIV activity (CVN-gp120 interaction) depends on the correct formation of disulfide bonds during recombinant production. Thus, inexpensive and functional production of rCVN requires an effective expression system which must be found among the bacteria, yeast, and transgenic plants, for the subsequent satisfying medical application. Moreover, the strong anti-HIV potential of CVN in trace concentrations (micromolar to picomolar) was reported for the in vitro and in vivo tests. Conclusion: To produce pharmaceutically effective CVN, we first need to identify the best expression system, with Escherichia coli, Pichia pastoris, Lactic acid bacteria and transgenic plants being possible candidates. For this reason, heterologous production of this valuable protein is a serious challenge. Since different obstacles influence clinical trials on microbicides in the field of HIV prevention, these items should be considered for evaluating the CVN activity in pre-clinical and clinical studies

Investigating the effects of Lactobacillus casei on some biochemical parameters in diabetic mice

Aims: Diabetes mellitus is a metabolic disorder characterised by inadequate  pancreatic insulin secretion or the insulin present being unable to perform its function properly. Consistent with the beneficial effects of probiotics and their ability to lower glucose levels, an impact on diabetes treatment is also expected. The aim of this study was to evaluate the effect of Lactobacillus casei on various either  biochemical parameters in a diabetic mice model.Methods: In the present study, 24 mice were divided into diabetic and control  groups. Further, each group was categorised into two subgroups. The diabetic and control subgroups were fed carrot juice or Lactobacillus casei in carrot juice. Diabetes was induced by streptozotocin (STZ). For 30 days, the mice were fed 2 ml carrot  juice, and Lactobacillus casei in carrot juice (with lactobacillus 109 cfu/ml) by gavage. Then, blood samples were collected to assay biochemical parameters.Results: The results of this study showed that Lactobacillus casei (ATCC39392) significantly reduced blood glucose (BG) levels in diabetic mice receiving the  probiotic, but did not cause a significant change in BG levels in control mice  receiving the probiotic. When comparing insulin, insulin-like growth factor I (IGF-I) and C-peptide in the four groups, it was found that insulin and C-peptide were  significantly different in all groups except for the control group treated with a mixture of probiotic Lactobacillus casei and carrot juice.Conclusion: Our results showed that probiotic Lactobacillus casei effectively  reduces BG levels in diabetic mice treated with this bacterium.Keywords: diabetes, Lactobacillus casei, probioti

Targeted delivery of silibinin via magnetic niosomal nanoparticles: potential application in treatment of colon cancer cells

Introduction: In recent years, various nanoparticles (NPs) have been discovered and synthesized for the targeted therapy of cancer cells. Targeted delivery increases the local concentration of therapeutics and minimizes side effects. Therefore, NPs-mediated targeted drug delivery systems have become a promising approach for the treatment of various cancers. As a result, in the current study, we aimed to design silibinin-loaded magnetic niosomes nanoparticles (MNNPs) and investigate their cytotoxicity property in colorectal cancer cell treatment.Methods: MNPs ferrofluids were prepared and encapsulated into niosomes (NIOs) by the thin film hydration method. Afterward, the morphology, size, and chemical structure of the synthesized MNNPs were evaluated using the TEM, DLS, and FT-IR techniques, respectively.Results and Discussion: The distribution number of MNNPs was obtained at about 50 nm and 70 nm with a surface charge of −19.0 mV by TEM and DLS analysis, respectively. Silibinin loading efficiency in NIOs was about 90%, and the drug release pattern showed a controlled release with a maximum amount of about 49% and 70%, within 4 h in pH = 7.4 and pH = 5.8, respectively. To investigate the cytotoxicity effect, HT-29 cells were treated with the various concentration of the drugs for 24 and 48 h and evaluated by the MTT as well as flow cytometry assays. Obtained results demonstrated promoted cell cytotoxicity of silibinin-loaded MNNPs (5-fold decrease in cell viability) compared to pure silibinin (3-fold decrease in cell viability) while had no significant cytotoxic effect on HEK-293 (normal cell line) cells, and the cellular uptake level of MNNPs by the HT-29 cell line was enhanced compared to the control group. In conclusion, silibinin-loaded MNNPs complex can be considered as an efficient treatment approach for colorectal cancer cells

Comparison of Inhibitory Effect of Curcumin Nanoparticles and Free Curcumin in Human Telomerase Reverse Transcriptase Gene Expression in Breast Cancer

Purpose: Telomerase is expressed in most cancers, including breast cancer. Curcumin, a polyphenolic compound that obtained from the herb of Curcuma longa, has many anticancer effects. But, its effect is low due to poor water solubility. In order to improve its solubility and drug delivery, we have utilized a β-cyclodextrin-curcumin inclusion complex. Methods: To evaluate cytotoxic effects of cyclodextrin-curcumin and free curcumin, MTT assay was done. Cells were treated with equal concentration of cyclodextrin-curcumin and free curcumin. Telomerase gene expression level in two groups was compared by Real-time PCR. Results: MTT assay demonstrated that β-cyclodextrin-curcumin enhanced curcumin delivery in T47D breast cancer cells. The level of telomerase gene expression in cells treated with cyclodextrin-curcumin was lower than that of cells treated with free curcumin (P=0.001). Conclusion: Results are suggesting that cyclodextrin-curcumin complex can be more effective than free curcumin in inhibition of telomerase expression

An overview on different strategies for the stemness maintenance of MSCs

Recent evidence suggests that mesenchymal stem cells (MSCs) have promising therapeutic potential for a broad range of diseases. Because the percentage of MSCs obtained from tissues is very low for cell therapy applications, ex vivo expansion of MSCs is necessary, but aging, loss of stemness and undesired differentiation of them during in vitro cultivation reduces their effectiveness. For achieving ideal therapeutic potential of MSCs in tissue regenerative purposes, it is necessary to retain their stemness properties in vitro. This review emphasis on the last updates in preserving the self-renewal capability of stem cells through in vitro expansion with different parameters

Comparative effects of Nucleostemin silencing in human Molt-4 and Jurkat leukemia T-ALL cells

     Nucleostemin (NS), a stem cell-abundant nucleolar protein, is critical for maintaining the self-renewal and proliferative properties of normal and cancerous stem cells. Recent data suggests that NS signaling is important for proliferation of T-cells and leukemia cells. This study was conducted to verify the role of NS in pathogenesis and treatment of T-cell acute lymphocytic leukemia (T-ALL). Our results revealed that RNA interference-mediated NS silencing primarily affected clonogenicproperty of T-ALL cells by limiting their self-renewal potential in vitro.These effects were accompanied with inhibition of proliferation and early apoptosis in Jurkat cells (p53-null) while late apoptosis in Molt-4 (p53 functional) T-ALL cells. Collectively, our results suggest that NS is a critical regulator in self-renewal and apoptosis of differentT-ALL cells. This suggests therapeutic potential of this gene in leukemia

Subcloning and expression of human alpha-fetoprotein gene in Pichia pastoris

Alpha-fetoprotein protein (AFP) is naturally found in fetal serum while production and appearance after birth is indicative of presence of malignant tumors. Therefore, by measuring this protein during fetal period and after birth, it is possible to diagnose abnormalities and tumors in fetus or the newborn. The objective of this study was to produce and purify AFP protein using DNA recombinant technology to apply in diagnostic kit preparations. In this study Pichia pastoris as metylotrophic yeast was used for AFP production. After construction of recombinant plasmid, pS1-AFP, electroporation and lithium chloride techniques were used for transferring to susceptible cells. The quantity and quality of the produced protein were checked by SDS-PAGE and ELISA methods. Selection of transformed mutant strains of Muts and culturing them in glycerol media (YPG) up to OD600=6 and their transfer to methanol media (YPM) with augmentation of methanol to 1% final concentration resulted in inducing protein production in auxotrophic media lacking histidine. This protein could be useful in monoclonal antibody production and in diagnostic kit preparations.Keywords: Alpha-fetoprotein, Pichia pastoris, cloning, expressio