جستجوی مقالات مرتبط با کلیدواژه "cell culture" در نشریات گروه "پزشکی"

Cell culture is a vital method in biological and biomedical research. The global cell culture market, valued at around USD 26.54 billion in 2023, is projected to surpass USD 63.60 billion by 2032. While two-dimensional cell culture has led to significant advancements in biology, its simplicity does not accurately reflect the complex in vivo environment. This can result in misleading data with limited predictive value for in vivo applications, prompting increased interest in three-dimensional (3D) cultivation methods. The 3D cell culture mimics the behavior and organization of cells in vivo by emulating the extracellular matrix (ECM), providing better insights into 3D interactions among cells and between cells and the matrix, thus reconstructing their natural microenvironment. In this review we will outline the various types of 3D models (include spheroids, organoids, bio-printed structures, and tissue chips). Subsequently, we will examine the methodologies employed to develop 3D culture systems (include four category methods). Lastly, the practical applications and challenges of these 3D models will be addressed. The future research will likely concentrate on incorporating cutting-edge technologies to improve the reproducibility and applicability of 3D models in research.

Ovine enzootic abortion (OEA) caused by Chlamyidia abortus is one of the most important abortive disease in small ruminants. Diagnosis of Ovine enzootic abortion depends on the isolation and detection of the agent or its nucleic acid. The aim of the present study was to detect Chlamydia abortus using both isolation method and real-time PCR in Brucella free flocks in Iran.

Twenty-eight vaginal and conjunctival swab samples which were Chlamydia abortus seropositive, were selected from ewes and does with recent abortion. Then the samples were tested by real-time PCR and positive molecular samples were inoculated into McCoy cells.

Using real-time PCR, 18 samples (64.3%) were positive and 7 (25%) of them were isolated in cell culture.

 The present results indicate that Chlamydia can play a relatively significant role in the abortion in does and ewes in Iran. Although the isolation of Chlamydia abortus have 100% specificity, because of low sensitivity, time consuming, cost and high probability of contamination, it is not suitable for routine laboratory diagnosis. Therefore, applying real-time PCR which have high sensitivity and specificity is recommended.

Human herpes virus type 1 (HSV-1) is a neurotropic pathogen that is infected more than 70% of the world population. The increasing of viral resistance to antiviral drugs and the emergence of side effects has motivated researchers to study the use of probiotics as new antiviral agents. The aim of the present study was to study for the first time the potential antiviral activity of Lactobacillus reuteri (L. reuteri) supernatant against HSV-1.

After measuring the cytotoxicity of L. reuteri supernatant by MTT assay, 1:16 dilution of it was added to HeLa cells before and after HSV-1 infection, after 1.5 hours incubation with HSV-1, and simultaneously with HSV- 1 infection. After 48 hours of incubation at 37°C, the viral titer and expression levels of UL54, UL52 and UL27 genes were measured by tissue culture infectious dose 50 (TCID ) and Real-Time PCR methods, respectively.

HSV-1 titer in the treatment conditions before infection, incubation with HSV-1, simultaneously with infection and after infection was reduced by 0.42, 3.42, 1.83, and 0.83 log 10 TCID /ml, respectively. When the bacterial supernatant was first incubated with the virus and then added to the cell, or when it was added simultaneously with the virus, the expression of the UL27, UL52, and UL54 genes decreased significantly (p<0.05). When the bacterial supernatant is added to the cell before or after virus infection, the expression of UL52 and UL54 genes does not change significantly (P>0.05).

The study findings indicated that the supernatant of L. reuteri has a potent anti-HSV-1 effect especially if it is incubated with the virus before inoculation into the cell. Its possible antiviral mechanism is to inhibit the virus by binding to it or changing the surface structure of the virus. Metabolites of L. reuteri can be considered as a novel inhibitor of HSV-1in- fection.

 Blastocystis sp. is an opportunistic worldwide protozoan that is life-threatening in immunocompromised individuals; several efforts were performed against Blastocystis sp. in in vivo and in vitro conditions to find an alternative for metronidazole (MTZ).

 The aim of the present study was to survey the cytotoxic effects of peptide mCM11 (modified CM11) and MTZ on ST1 and ST3 of Blastocystis sp. in vitro.

 In the present study, after the culture of the parasite (in Dulbecco’s modified Eagle medium [DMEM] and Caco-2 cell lines), the anti-parasitic effect of different concentrations of either MTZ (1.25-80 μg/mL) or mCM11 (1-128 μg/mL) for 24, 48, and 72 hours was evaluated via a 3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide assay and flow cytometry.

 A dose-time-dependent decrease in cell viability was detected after cell line and parasite exposure. The cytotoxic effect of the peptide on cell lines increased during 24, 48, and 72 hours, and the greatest anti-parasitic effect was observed at the highest concentration (128) after 72 hours. In addition, the ST3 showed more sensitivity to the mCM11.

 The findings of the current research about the mCM11 peptide were promising compared to MTZ, and by further optimizing the peptide, better effects can be imagined for it, so that it can be a potential alternative to MTZ. However, more studies with a case-control design seem to be necessary in in vivo conditions.

 The exploration of bryophytes biodiversity in Indonesia due to its abundance and the bioactivity of its phytochemical content, such as alkaloids and polyphenols, has received increased interest. Despite some species proven to possess pharmacological properties, the antiproliferative study of Indonesian native moss, such as the Pogonatum genus, is limited. Hence, this study aims to evaluate the anticancer effects of Pogonatum neesii Dozy antiproliferative activity on colon and cervical cancer through in silico and in vitro methods.

 Molecular docking analysis using Autodock VINA in PyRx softwre was conducted between natural compounds found on P. neesii and several target proteins, DNA (cytosine-5)- methyltransferase 1 (DMT-1) (Protein Data Bank (PDB) id: 4WXX) in colon cancer and B-cell lymphoma 2 (Bcl-2) (PDB id: 4LXD) in cervical cancer. Afterwards, total phenolic and alkaloid contents were measured. Subsequently, P. neesii was tested on HaCaT (keratinocytes), HEK293 (human embryonic kidney), HT-29 (colorectal cancer models) and HeLa (cervical cancer model) to observe its cytotoxicity.

 Out of eight compounds, chlorogenate was found to exert the best binding energy with target proteins, although it had lower binding affinity than the protein’s natural ligand. However, the biological, drug-likeness, and toxicity analysis suggested the drug potency of the compound, thus we did the in vitro analysis. P. neesii showed significant cytotoxic effects on HT-29 and HeLa cells, while it did not exert any cytotoxic effects on HaCaT and HEK-293 cells, at the same concentrations.

 P. neesii has been shown to have the potential as an anticancer agent through in silico and in vitro analysis, where the extract showed selective cytotoxicity towards cancer cell lines and cytocompatibility towards normal cell lines. Chlorogenate was pinpointed as the compound with the most activity and interaction with the target proteins in both cancers.

Chemotherapy drugs used to treat lung cancer are associated with drug resistance and severe side effects. There have been rising demands for new therapeutic candidates and novel approaches, including combination therapy. Here, we aimed to investigate the combinatorial effect of a dendrosomal formulation of curcumin (DNC) and daunorubicin (DNR) on the A549 lung cancer cell line.

We performed cytotoxicity, apoptosis, cell migration, colony-formation capacity, and gene expression analysis to interpret the mechanism of action for a combination of DNC and DNR on A549 cells.

Our results revealed that the combination of DNC and DNR could synergistically inhibit the A549 cells’ growth. This synergistic cytotoxicity was further approved by flow cytometry, migration assessment, colony-forming capacity and gene expression analysis. DNR combination with DNC resulted in increased apoptosis to necrosis ratio compared to DNR alone. In addition, the migration and colony-forming capacity were at the minimal range when DNC was combined with DNR. Combined treatment decreased the expression level of MDR-1, hTERT and Bcl-2 genes significantly. In addition, the ratio of Bax/Bcl2 gene expression significantly increased. Our analysis by free curcumin, dendrosomes and DNC also showed that dendrosomes do not have any significant cytotoxic effect on the A549 cells, suggesting that this carrier has a high potential for enhancing the curcumin’s biological effects.

Our observations suggest that the DNC formulation of curcumin synergistically enhances the antineoplastic effect of DNR on the A549 cell line through the modulation of apoptosis/necrosis ratio, as well as Bax/Bcl2 ratio, MDR-1 and hTERT gene expression.

Neural stem cells (NSCs) are multipotent stem cells residing in the central nervous system that is capable of self-renewal to support ongoing requirements for neurogenesis in the adult brain. Since NSCs are considered potential candidate cells for neuro-regenerative medicine, applying safe induction methods for them is very important. Synthetic modified-mRNA (mmRNA) as an alternative to traditional DNA- or protein-based methods, is regarded as a powerful tool for inducing short-term gene expression in cells with no genetic manipulation.

Here, we aimed to develop an optimized condition for mmRNA transfection in primary NSCs. In vitro-transcribed EGFP mmRNA (mmRNAEGFP) was delivered to human embryonic kidney cells (HEK293T) and mouse NSCs by using two commercial agents, Lipofectamine-2000 (LF2000) and TransIT. Also, a plasmid DNA was used to transfect cells considered EGFP-expressing positive control. In addition, the poly(A) tail (poly adenosine tail) elongation and chloroquine (CQ) treatment were performed to improve transfection efficiency. Finally, flow cytometry, fluorescence microscopy, and MTT assays were performed to assess the cells.

In comparison with HEK293T, NSCs were very sensitive to transfection, the efficacy of transfection using DNA/LF2000 was higher in HEK293T cells, but mmRNAEGFP/ TransIT showed better transfection efficacy in NSCs. Poly(A) tail elongation; also, treating the cells with CQ before transfection significantly improved its efficacy.

The mmRNA poly(A) tail elongation and the use of specific transfection agents in combination with TLR inhibitors can lead to a more effective transfection in NSCs.

Cell culture has a crucial role in many applications in biotechnology. The production of vaccines, recombinant proteins, tissue engineering, and stem cell therapy all need cell culture. Most of these activities needed adherent cells to move, which should be trypsinized several times until received on a large scale. Although trypsin is manufactured from the bovine or porcine pancreas, the problem of contamination by unwanted animal proteins, unwanted immune reactions, or contamination to pathogen reagents is the main problem.

This study investigated microbial proteases as a safe alternative for trypsin replacement in cell culture experiments for the detachment of adherent cells.

The bacteria were isolated from the leather industry effluent based on their protease enzymes. After sequencing their 16S ribosomal deoxyribonucleic acid, their protease enzymes were purified, and their enzyme activities were assayed. The alteration of enzymatic activities using different substrates and the effect of substrate concentrations on enzyme activities were determined. The purified proteases were evaluated for cell detachment in the L929 fibroblast cells compared to trypsin. The separated cells were cultured again, and cell proliferation was determined by the MTT assay.

The results showed that the isolated bacteria were Bacillus pumilus, Stenotrophomonas sp., Klebsiella aerogenes, Stenotrophomonasmaltophilia, and Bacillus licheniformis. Among the isolated bacteria, the highest and the lowest protease activity belonged to Stenotrophomonas sp. and Klebsiella aerogenes, with 60.34 and 11.09 U/mL protease activity, respectively. All the isolated microbial proteases successfully affected L929 fibroblast cells’ surface proteins and detached the cells. A significant induction in cell proliferation was observed in the cells treated with Klebsiella aerogenes protease and Bacillus pumilus protease, respectively (P < 0.05).

The obtained results suggested that microbial proteases can be used as safe and efficient alternatives to trypsin in cell culture in biopharmaceutical applications.

Although investigating the probable side effects of post intraoperative radiotherapy wound fluid secretion (PIWFS) is crucial, especially in clinical cases, no report has been published on the effect of PIWFS on the remaining tumor cells (in the vital state) in cavity side margins or surrounding regions. These tumor cells might be directly/indirectly exposed to intraoperative radiation therapy (IORT). Here, for the first time, we investigated the effect of PIWFS on tumor cells of the same patient extracted from the excised tumor in the spheroid form.

We generated 8 human-derived breast tumor spheroids from 4 patient specimens who received to IORT, dissociated and cultured them in microfluidic devices. The spheroids from each sample were treated with the patients’ PIWFS and DMEM medium separately. Two different parameters, called area and number of detached cells (NDCs), were determined and investigated to evaluate the spheroids’ vital and proliferative states.

The results showed severe transformation in tumor spheroids’ function into more invasive and proliferative functions after treatment with PIWFS.

Although the radiation-induced bystander effect may have a role in this observation, further experiments must be done to better clarify the probable desired or non-desired effects of post-IORT secretion for both the remaining tumor cells and the surrounding immune cells.

Metal nanoparticles may apply to the cavity walls as acceptable antibacterial agents. The present work was conducted to assess the effect of silver nanoparticles (NPs) on the resin cements Shear Bond Strength (SBS) using one self-etch and one etch and rinse resin cement.

Sixty intact noncarious extracted, human third molars were selected. All teeth were cut out in the middle of dentin thickness. Resin cement samples were randomly assigned to five groups: (A) Duolink cement without any further surface treatment; (B) Duolink cement with silver NPs (0.5%) dentin surface treatment after acid etching and before the One Step Plus (OSP) adhesive system; (C) Duolink cement with silver NPs (0.5%) dentin surface treatment before acid etching and OSP; (D) Panavia F2.0 cement without any further surface treatment; and (E) Panavia F2.0 cement with silver NPs (0.5%) dentin surface treatment. SBS was assessed by using universal testing machine. Then, human gingival fibroblasts (HGF) cells were treated with tested materials. The biocompatibility of resin cement was evaluated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay.

The maximum and minimum of the SBS at the presence of silver NPs were observed in group E (15.81±0.91, P-value<0.01) and group B (12.01±0.15, P-value <0.05), respectively. In comparison with the controls, the resin cements incorporated  with 0, 0.2% and 0.5% silver NPs, 20.34%, 22.00% and 22.67% for Panavia F2.0 and 36.84%, 37.34% and 38.17% for Duolink flow - decreased in total cell number, respectively (P-value<0.01).

These findings demonstrated that Panavia F2.0 cement with silver NPs dentin surface treatment showed satisfactory result in the SBS compared to Panavia F2.0 cement without any further surface treatment.

Given the prevalence of fertility problems in couples and the defect in embryo implantation as well as the low success rate of assisted reproductive techniques, it is necessary to investigate the causes of this phenomenon. Type 2 diabetes mellitus (T2DM) is a metabolic disease with multiple effects on various organs as well as the endometrium. In this study, the effects of endometrial cell culture on the expression of α3 and β1 integrin genes and protein in type 2 diabetic rats were investigated.

In this experimental study, 35 female rats were divided into five groups: control, sham, diabetic, Pioglitazone-treated and Metformin-treated groups. First, rats were maintained in diabetic condition for 4 weeks. Then, treatment was performed for the next four weeks. Four weeks after induction of diabetes, rats were sacrificed at the time of embryo implantation. The uterus was removed. Endometrial cells were isolated and cultured for 7 days. Immunocytochemistry staining was used to confirm endometrial cells. Expression of α3 and β1 integrin genes was determined by real-time polymerase chain reaction (PCR) technique and the α3β1 protein content measured using Western blot both before and after endometrial cell culture.

The expression level of α3 integrin gene in the Pioglitazone-treated group compared with metformin-treated group was significantly decreased (P<0.001). The same result was observed in β1 integrin gene expression (P=0.004). Also, the α3β1 protein level increased in all diabetic groups, but its reduction was significantly greater in pioglitazonetreated group (P=0.004)

T2DM altered the expression of α3 and β1 integrin genes and related proteins, which endometrial cell culture regulated this disorder. According to these results, may be the endometrial cell culture can reduce the adverse effects of diabetes on α3 and β1 integrin expression at the level of gene and protein, in endometrial cells.

Spirulina (Arthrospira) exerts a wide spectrum of pharmacological activities that are largely attributed to its phycobiliprotein content,  mainly  to  C-phycocyanin.  C-Phycocyanin  is  a  natural  blue  pigment  with  many commercial  applications  in  foods, cosmetics, and medicines.  In this study, the stimulatory effect of  C-phycocyanin on the immune system was investigated using blood tissue and Peripheral  Blood  Mononuclear  Cells  (PBMC)  and  the  measurement  of  inflammatory  cytokine,  interferon-gamma, which  has important effects on dedicated immune responses. For this purpose, extraction of PBMC blood cells from blood tissue The range of  purified  C-phycocyanin  extract  concentrations  with  drug  puritywas:  1  µg/ml,  10  µg/ml,  100  µg/ml,  250  µg/ml.  Then, the response of PBMC cells to C-phycocyanin at the protein level was investigated. Finally, interferon-gamma was measured using the culture supernatant and ELISA sandwich method.  Descriptive  analysis  of  the  read  concentrations  results  by  ELISA  technique  showed  that  C-phycocyanin  is dosedependent  and  the  results  of  the  effect  of  C-phycocyanin  on  PBMC  cells  in  blood  tissue  showed  the strengthening  of  the immune system by increasing the amount of inflammatory cytokines. According to the results of the analysis of variance, it isobserved that the p-value is less than 0.05. This means there is a significant difference between the mean read concentration of ELISA in different concentrations of cytokines. The results of the experiments demonstrate that C-phycocyanin activates PBMC cells in a manner that is consistent with the recruitment of diverse populations of leukocytes in response to inflammatory and infectious signals.

Rabies is a zoonotic fatal viral disease caused by the rabies virus of the genus Lyssavirus, and the family Rhabdoviridae. Challenge virus standard (CVS-11) strain of rabies virus is a key element in rabies reference laboratories, as many gold-standard tests depend on a suitable titer of this strain for interpretation of results. The present study investigated the optimal CVS-11 production in BSR cells (a clone of BHK-21).

We analyzed the kinetic growth of BSR cells in a T-flask and inoculated BSR cells with different MOI of CVS-11 strain of rabies virus, and harvested the produced virus at different time points. 

Our data showed that BSR cells had a doubling time of around 24-30 h, and at least 95% of cells kept their viability three days after culture. The virus reached the highest titer when the cells were infected at an MOI of 0.1 in DMEM medium, equal to 1.5 × 107 fluorescent focus units (FFU)/ml. Time-course analysis of CVS-11 titer showed that the highest titer was achieved around 72 h post-infection. All tests were performed in triplicate.

Since producing the virus in mammalian cell culture is an expensive and complicated method, optimizing the virus production process may be an excellent strategy to lower the cost, save the laboratory resources and maximize productivity.

Cell experiments are vitally dependent on CO2 incubators. The heating system of usual incubators result in undesirable induction of Electromagnetic (EM) fields on cells that result in decreased accuracy in bio-electromagnetic tests. EM shields can cause a considerable decrease in the stray fields and eliminate the undesirable induction.

CST-2019 is used for simulations. five different shielding systems have been examined in this paper. We try to modify shape and material used for shielding to achieve better result. (Iron, Mu-Metal, steel).

We introduce a simple practical design, together with variations of previously reported ones, and numerical evaluation of their magnetic field attenuation.

The targeted design decreases the field within the shield to about 0.03 times of the incident magnetic field, while having holes for air and CO2 exchange.

Taxol (Paclitaxel) has been highly recommended to control and treat a wide range of cancers. Taxus baccata L. is primary and natural origin of Taxol.

Due to severe restriction and prohibition of cutting T. baccata, the alternative way for Taxol production is to apply in vitro culture method which was implemented at the current study. Here, for the first time, the effect of amino acid complex as elicitor along with chitosan in cell suspension culture ofT. baccata for enhancing Taxol production was studied.

Young leaves of T. baccata as explants were cultured in different callus induction treatments. At the next step, the calli from the best callus induction treatment were transferred to cell suspension cultures containing chitosan and amino acid complex alone or in combination with each other. Taxol content in treatments were measured by HPLC.

MS medium supplemented with 2,4-D at 2 mg/L and NAA at 1 mg/L proved to be the best treatment of callus induction (100 %), fresh weight (495 mg) and dry weight (272 mg) of calli. Also, HPLC analysis confirmed the maximum production of Taxol (1.96 mg/g) in MS medium having 2 ml/L amino acids complex with 10 mg/L chitosan.

Applying amino acid complex as elicitor with chitosan is suggested for enhancing Taxol production in cell suspension culture of T. baccata.