جستجوی مقالات مرتبط با کلیدواژه "flow cytometry" در نشریات گروه "پزشکی"

Viable but non-culturable (VBNC) is a specific physiological state in which living bacteria lose the ability to grow and form colonies on conventional bacterial growth media; however, these cells remain metabolically active.

Considering the importance of the pathogenicity of Shigella flexneri and the possibility of its transmission through contaminated water and food, we aimed to study the possibility of this bacterium entering the VBNC state under osmotic and nutritional stresses at low temperature (4°C).

Shigella flexneri was inoculated in distilled water and NaCl solutions with concentrations of 0, 5, 10, 15, 20, 25, and 30% at 4°C. The cultivability of the bacteria was checked daily. After observing the loss of cultivability, entry into the VBNC state was assessed using RT-PCR, flow cytometry, and fluorescence microscopy.

After cultivation of S. flexneri in different concentrations of salt solution, the time of non-cultivability was observed at concentrations of 0, 5, 10, 15, 20, 25, and 30%, respectively, on days 19, 20, 21, 22, 22, 24, and 24. Expression results of ipaH and ipaD genes using RT-PCR, along with the presence of live cells detected by flow cytometry and fluorescent staining, indicated that the cells had entered the VBNC state.

Shigella flexneri can survive and enter the VBNC state under nutritional and osmotic stresses at low temperatures (4°C). In this state, although the bacterium is alive, it cannot be detected by conventional culture-dependent methods. Due to the potential risk of bacterial recovery, conventional techniques may be insufficient to detect the presence of pathogens in the VBNC state when evaluating water and food.

Several long non-coding RNAs are implicated in the development and metastasis of cancer. The non-coding RNA HOTAIR is known to play a significant role in the progression of various cancers.

This study aimed to assess the impact of HOTAIR dysregulation on stemness markers in AGS and MKN45 gastric cancer cell lines.

Downregulation and upregulation of HOTAIR were induced using small interfering RNA (siRNA) and a HOTAIR overexpression vector, respectively. The surface stemness markers CD24 and CD44 were analyzed using flow cytometry. Additionally, the effects on the expression of two stemness markers, NANOG and P21, were investigated using Quantitative Reverse Transcription PCR (qRT-PCR).

Flow cytometry analysis showed that HOTAIR modulates the cell-surface expression of CD44 and CD24 in gastric cancer cells. Furthermore, dysregulation of HOTAIR significantly affected the expression of the genes P21 and NANOG.

Our studies suggest that HOTAIR may regulate the stemness characteristics of gastric cancer cell lines.

Radiotherapy, a highly effective method of radiation-based treating cancers, can reduce the size of tumors and affect healthy tissues. Radiation-induced lymphopenia as a side effect of radiation therapy can reduce the effectiveness of the treatment.
This study aimed to examine how taurine can protect peripheral blood lymphocytes from radiation-based apoptosis.
In this experimental study, the effects of the taurine on lymphocytes were studied, and blood samples were divided into three groups: a negative control group that was not treated, a positive control group that was treated with cysteine (100 μg/ml), and a group that was treated with taurine (100 µg. mL-1) in three different doses (4, 8 & 12 Gy) before irradiation. The percentage of apoptotic and necrotic lymphocytes was measured using flow cytometry 48 and 72 hours after the irradiation, respectively.
According to the groups treated with taurine, the number of lymphocytes undergoing apoptosis was lower and higher compared to the negative and positive control groups, respectively. The decrease in this value was more pronounced 48 hours after radiation compared to 72 hours. Furthermore, there was a slight increase in the number of apoptotic lymphocytes with increasing radiation dose. 
Taurine effectively protects human peripheral blood lymphocytes from radiation-based apoptosis.

Systemic sclerosis (SSc) is a chronic autoimmune disorder characterized not only by fibrosis and vasculopathy but also by inflammation. Previous studies have demonstrated monocyte involvement in SSc development, suggesting a role for immune dysfunction in SSc pathogenesis. To investigate the relationship between SSc’s clinical manifestations and altered levels of monocyte subpopulations. Twenty-six patients meeting the ACR/EULAR SSc criteria along with twenty healthy individuals as the control group, were enrolled in the study. Peripheral blood mononuclear cells (PBMCs) were obtained from heparinized blood samples of both the SSc patients and the control group. Subpopulations of monocytes were assessed based on HLA-DR, CD14, and CD16 expression using multi-color flow cytometry. The one-way ANOVA, Student’s t-test, and Mann-Whitney U test were employed for normally and non-normally distributed data. The Spearman correlation test was utilized to identify correlations between the variables. The SSc patients showed a significant increase in the number of circulating peripheral blood monocytes (p<0.001). The percentage of CD16+ monocyte subpopulations was higher in the SSc cases compared to the control group. A significant decrease in the ratio of classic to non-classic monocytes was observed in SSc cases (7.43%) compared to the control group (52.09%, p<0.001). No association was observed between monocyte subpopulations and clinical characteristics of SSC. Our results showed an increase in the level of CD16+ monocytes in patients with SSc compared to healthy individuals. Further investigation is required to determine the clinical significance of this alteration.

Peptides from lactic acid bacteria provide health benefits and can inhibit the growth of pathogenic organisms. The present work aimed to isolate and characterize peptides with antibacterial activity from Lactobacillus plantarum 1407.

Peptides were isolated and purified from L. plantarum 1407. The effect of various physiological parameters on the antibacterial activity of the isolated peptides was analyzed. The mode of action of the peptides on indicator organisms was observed using transmission microscopy analysis and flow cytometry analysis .

Antibacterial activity and mode of action of peptides isolated from L. plantarum 1407 against gram-positive and gram-negative bacteria have been studied. L. plantarum culture exhibited maximum antibacterial activity at 40 °C, pH 8, and 0.7% salt concentration. The cell-free supernatant (CFS) was concentrated using a 3 kDa ultrafiltration membrane and the peptide fractions (<3 kDa) were further fractionated using Sephadex G-25 gel filtration chromatography. The antibacterial activity of the eluted fractions (F1 to F4) was evaluated using flow cytometry and transmission electron microscopy. F3 fraction exhibited increased antibacterial activity than F1, F2, and F4 fractions against the indicator organisms. Cell membrane damage and leakage of cytoplasmic content of the bacterial cells treated with the antibacterial F3 fraction peptides were observed.

The active peptides from L. plantarum 1407 can be potentially used for the treatment of bacterial infections.

Breast cancer (BC) is the second leading cause of death due to cancer among women worldwide. Therefore, the present study investigates the cytotoxic effects of piperine on the breast cancer cell line (MCF7) and the genes of the apoptotic pathway. 

This research was performed to assess the effect of piperine on BC cells and the change in the expression level of bax gene through the induction of apoptosis.

MCF-7 cells were prepared by the Pasteur Institute, Iran. Cytotoxicity of piperine at concentrations of (5, 10, 15, 20, and 25 µM) during 24, 48, and 72 hours was evaluated by MTT assay. The cell apoptosis and bax gene expression were evaluated by Flow Cytometry and qReal-time PCR, respectively. Finally, the statistical analysis of MTT and RT- PCR data was done by SPSS software version 22. 

The piperine showed concentration-dependent cytotoxic effects on the MCF-7 cell line in MTT assay. The bax gene expression level has a significant increase in piperine-treated cells compared to the untreated ones. The MCF-7 cell apoptosis at IC50 concentration of piperine was measured at 58.3% during 48-h treatment. 

In general, it can be concluded that piperine has cytotoxic effects against breast cancer by inducing apoptosis via overexpressing of bax.

Galangin (3,5,7 tri-hydroxy flavone), naturally active flavonoid is derived from the rhizomes of Alpinia officinarum is proven to be effective antioxidant, anti-inflammatory, anticancer. However, protective effects of galangin in chronic renal failure (CRF) is not explored.

This study is aimed to investigate the nephroprotective activity of galangin in adenineinduced CRF rats.

Adenine induced rats were administered galangin 20 mg/kg and 40 mg/kg body weight (BW) serum renal and hepatic parameters, and renal antioxidant-lipid peroxidation parameters, histological studies were carried out. The mechanism of action was investigated by flow cytometry.

Galangin has normalized serum renal and hepatic parameters, reduced oxidative stress and lipid peroxidation. Histopathology confirms nephroprotection. The percentage number of cells expressing transforming growth factor beta (TGF-β) was reduced with galangin treatment.

Galangin exerts nephroprotection in adenine induced CRF by inhibiting TGF-β expression.

Acute lymphoblastic leukemia (ALL) is the most prevalent childhood cancer. ALL is a heterogeneous type of malignancy and treatment protocols vary based on the immunological classification of ALL. The critical step for treating ALL is to identify immunological subgroups by flow cytometry findings. 

In this study, immunophenotypic information was evaluated for the first time in children with ALL in Qazvin City, Iran.

This cross-sectional study reviewed the clinical and laboratory data of children with ALL during 2019-2020. Next, children with ALL were immunophenotyped by flow cytometry applying a panel of the specific monoclonal antibodies for some clusters of differentiation (CD) molecules, including CD20, CD21, CD10, CD34, CD38, and terminal deoxynucleotidyl transferase (TdT). The data were separately analyzed using SPSS software, version 24.

Of 52 children with ALL in the age range of 6 months to 15 years, 23 children (44.23%) had B-ALL-ProB (pro-B) cell immunotyping features, 26 (50%) had B-ALL-PreB (pre-B) cell immunotyping features, and 3 (5.7%) had T-cell immunotyping features. The ages of T-cell group children were higher than those of B-cell group children. The most common clinical and laboratory findings were fever (26 cases, 55.31%). In 55% of children, periodic acid-schiff (PAS) staining was positive. The presence of the terminal deoxynucleotidyl transferase (TdT) enzyme was higher in B-cell patients than in T-cell cases. Children with CD34+ were higher in the pro-B group than in the pre-B group. 

our study shows that the immunophenotypic characteristics of children with ALL are more similar to previous reports and can be used for monitoring and prognosis of children with ALL in Qazvin City.

 More than 768 million people have been affected by COVID-19. Identifying lymphocyte subsets and cytokine level abnormalities in COVID-19 patients is essential to gain new insights and data on immunity mechanisms against viral infections.

 We used flow cytometry to determine the relationship between disease severity, lymphocyte subsets distribution, and cytokine level alterations in COVID-19 patients.

 Totally 94 COVID-19 patients (32 mild, 31 moderate, and 31 severe) and 27 healthy individuals were included in the cross-sectional study. The distribution of peripheral lymphocyte subsets and cytokine levels was assessed by flow cytometry.

 The percentages of CD56+ Natural Killer (NK) cells in all patient groups and total T lymphocytes in moderate and severe groups were significantly lower than those in the control group (P < 0.001). Also, IL-2 (P < 0.001), IL-17A (P < 0.001), IL-4 (P < 0.001), IL-6 (P < 0.001), TNF-α (P = 0.004), IP-10 (P < 0.001), IFN-λ1 (IL-29) (P < 0.001), IFN-λ2/3 (IL-28A/B) (P = 0.011), IFN-β (P < 0.001), IL-10 (P < 0.001), and IFN-γ (P < 0.001) levels were statistically higher in patients than in the controls.

 Our data revealed that increased levels of certain cytokines in peripheral blood contribute to disease severity. Increased CRP (OR: 1.012, %95 CI: 1.002 - 1.023, P = 0.038) and IL-10 (OR: 1.068, %95 CI: 1.000 - 1.141, P = 0.049) levels, decreased CD56+ NK percentage (OR: 0.576, %95 CI: 0.376 - 0.882, P = 0.011) and lymphocyte count (OR: 0.02, %95 CI: 0.001 - 0.368, P = 0.009), and the presence of diabetes mellitus and mechanical ventilation were independent predictors of mortality.

CD39 is an inhibitory checkpoint exerting rate-limiting effects on the ATP-adenosine pathway. It can be targeted to block adenosine-mediated immunosuppression. To analyze the relationship between the CD39 expression and clinicopathological characteristics including FIGO stage, lymph node and distant metastasis, and to further explore its potential role in cervical cancer. Peripheral blood was collected from 59 healthy people and 43 patients with cervical cancer. The percentage and absolute counts of CD3-positive, CD4-positive and CD8-positive T lymphocytes, CD4/CD8 ratio and the percentage of the CD39+ T cells in T lymphocytes were assessed by flow cytometry, and their correlations with clinical parameters were analyzed. Absolute numbers of CD8+ T lymphocytes, CD4/CD8 ratios, and the percentage of the CD39+ T cells were linked with FIGO stage, lymph node metastasis, and distant metastasis. The total numbers of CD8+ T lymphocytes were significantly higher in the peripheral blood of patients with cervical cancer in the early and middle stages than in the advanced stage. In addition, patients with early and middle-stage cervical cancer had considerably lower percentage of CD4+ CD39 + and CD8 + CD39 + T lymphocytes than those with advanced cervical cancer. These results suggest that the absolute counts of CD8+ T lymphocytes may be associated with the patient’s prognosis and that the CD39 molecule, expressed on the surface of CD8+ T cells, is also related to FIGO stage, lymph node metastasis, and distant metastasis. CD39 expression on CD8-positive T cells exhibits a negative correlation with the number of CD8-positive T lymphocytes.

Antigen presentation using bacterial surface display systems, on one hand, has the benefits of bacterial carriers, including low-cost production and ease of manipulation. On the other hand, the bacteria can help in stimulating the immune system as an adjuvant. For example, using bacterial surface display technology, we developed a hepatitis C virus (HCV) multiple antigens displaying bacteria's surface and then turned it into a bacterial ghost.

The HCV core and NS3 proteins' conserved epitopes were cloned into the AIDA gene plasmid as an auto transporter. The recombinant plasmid was then transformed into Escherichia coli (E. coli) Bl21 (DE3). Recombinant bacteria were then turned into a bacterial ghost, an empty cell envelope. Whole-cell ELISA, flow cytometry, and Western blot techniques were used for monitoring the expression of proteins on the surface of bacteria.

A fusion protein of HCV core-NS3-AIDA was successfully expressed on the E. coli Bl21 (DE3) surface and confirmed by western blotting, Enzyme-Linked Immunosorbent Assay (ELISA), and flow cytometry detection techniques.

The presence of HCV antigens on non-pathogen bacteria surfaces holds promise for developing safe and cost-benefit-accessible vaccines with optimal intrinsic adjuvant effects and exposure of heterologous antigens to the immune system.

Fibrosing pneumonia (FP) is classified into usual interstitial pneumonia (UIP) and nonspecific interstitial pneumonia (NSIP), each having its own etiology and prognosis. Both types of FP are progressive and chronic conditions with distinct etiologies. Cytokines and inflammatory mediators play critical roles in the pathogenesis of FP. Among them, the role of transforming growth factor beta-1 (TGF-β1) and modulators triggering fibrosis are not well understood. In this study, the expression of triggering receptor expressed on myeloid cells-1 (TREM-1) as a stimulator for the production of TGF-β1 and also CD4+CD25+Foxp3+ regulatory cells were investigted in FP patients. Sixteen UIP, 14 NSIP and 4 pulmonary fibrosis following Mycobacterium tuberculosis (TB) infection patients, were compared with 12 healthy controls. The frequency of blood CD14+TGF-β1+ and CD14+TREM1+-gated monocytes and CD4+CD25+Foxp3+ regulatory T cells (Treg), as well as the plasma levels of TGF-β1 and IL‑10 were measured. Fibrosis patients compared to healthy controls had a greater frequency of CD14+TGF-β1+ [15.9 (0.2-88.2) vs. 0.6 (0.2-11.0)] and CD14+TREM1+ [21.1 (2.3-91.2) vs. 10.3 (3.1-28.6)]-gated monocytes, and CD4+CD25+Foxp3+ [1.2 (0.3-3.6) vs. 0.2 (0.1-0.4)]-gated lymphocytes. Plasma TGF-β1 were also significantly increased in patients with fibrosis compared to healthy controls [9316.2 (±5554.4) vs. 3787.5 (±2255.6)]. These results confirm the importance of TGF-β1 and TREM1 in pulmonary fibrosis. It seems that this reciprocal cycle in healthy people is modulated by the production of IL‑10 by Treg cells, thus limiting fibrosis, as observed in patients following TB infection. Further investigations are recommended to evaluate possible immunomodulatory mechanisms defects in pulmonary fibrosis.

Diagnosing hematologic malignancies requires implementing several tests. This study aims to evaluate the chromosomal changes in patients with myeloid disorders and compare the results of flow cytometry and cytogenetics with the initial diagnosis performed by the oncologist.

115 patients with myeloid disorders, 57.2% males and 42.8% females with a mean age of 50.3 years, previously diagnosed by an oncologist based on the clinical features, complete blood count, and peripheral blood smear interpretations, were considered. Moreover, flow cytometry and cytogenetic analysis were implemented on the bone marrow samples.

Cytogenetic results showed that 30% of patients with myeloid disorders had abnormal karyotypes. 77% of patients with myelodysplastic syndromes, 65% of acute myeloid leukemia, and 30.7% of chronic myeloid leukaemia indices showed normal karyotypes, and the others resulted in common and uncommon abnormalities, including the translocation (13;17), 92, XXYY, and del (4q). Considering the flow cytometry and karyotype results, the improved diagnoses were made for 41 patients who had not been diagnosed initially.

This study showed that, in some cases, an initial diagnosis is inconsistent with the flow cytometry and karyotype analysis results. Also, the flow cytometry results may differ from the karyotype depending on the case. Therefore, combining the results obtained by the cytogenetic investigation, flow cytometry, fluorescence in situ hybridization, and molecular testing is preferable to provide a comprehensive report for the appropriate disease diagnosis and prognosis.

Differential diagnosis of chronic lymphoproliferative disorders (CLDs) has remained challenging due to the highly variable morphology features and immunophenotyping. Currently, the development of multiple-marker panel analyses by flow cytometry has opened a broad way for diagnosis of CLDs.

We analyzed the peripheral blood and bone marrow samples of 131 patients with B-cell CLDs (including 91 chronic lymphocytic leukemia (CLL), 15 atypical CLL, 14 mantle cell lymphoma (MCL), and 11 CD5-/CD10-lymphoma patients) from April 2018 to April 2019, using a panel of specific markers by flow cytometry.

Our results indicated that the expression pattern of CD22, CD23, FMC-7, and CD5 allowed us to accurately and differentially diagnose the B-CLL, MCL, and CD5-/CD10- lymphoma, while it was not capable of differentiating MCL from atypical CLL. We, however, found that the expression patterns of CD38 and immunoglobulin light chain differed significantly between atypical B-CLL and MCL. CD38 and lambda light chain were remarkably expressed in MCL patients (92.8% and 85%, respectively) compared to the atypical CLL (1.1% and 0% respectively), with the p value less than 0.001 for both markers. In contrast to MCL patients, all the patients with atypical CLL, expressed kappa light chain. The immunohistochemistry method used for cyclin D1 confirmed that the flow cytometry detection of kappa and lambda light chains could provide a new approach with high sensitivity (91%) and moderate specificity (50%) to distinguish MCL patients from atypical B-CLL.

Expression of CD5, CD20 (bright), CD22, FMC-7, CD38, and lambda light chain with no expression of CD23 can accurately detect MCL and differentiate it from atypical B-CLL.

One of the well-accepted beliefs about natural products, considering the advances of recently appearing new edges and features of herbal medicine, is paying more attention to cancer treatments. However, they have not been properly studied with reasonable/reliable clinical trials in human subjects in most cases. Therefore, seeking in vitro effects of herbs like Melissa officinalis (MO) in cancer therapy to identify the involved possible mechanism in conjugation with configurative/morphological aspects of treated cells seems quite necessary. In this study, we evaluated the co-treatment effect of anti-cancer drug methotrexate (MTX) and MO on HeLa cancer cells.

MTT assay was applied to assess the quantitative cytotoxicity effect of both MTX and Mo. Apoptosis assay via flow cytometry was used to determine the amount of apoptotic and necrotic cells. To further investigate the anti-cancer effects, DAPI staining and DNA ladder assays are used qualitatively to detect changes in the nuclei of cells that are a sign of apoptosis occurring and morphological modifications of DNA.

MTX and MO mixture showed high cytotoxicity and apoptosis rate compared to untreated cells. Furthermore, the morphological changes of MTX and MO mixture were more evident than that of single MO, MTX, and control groups.

These data regarding cell growth reduction and apoptosis induction in HeLa cancer cells showed that MTX and MO mixture can be an appropriate platform for cancer therapy.

Breast cancer (BC) is the most common cancer in women, and its prevalence has increased dramatically in recent years. Many treatments for BC have been proven, one of which is the utilization of natural products and herbal derivatives. Among natural plants, plants of the Apiaceae family like Eryngium have been studied, which entail antioxidant, antimicrobial, and most importantly, anti-cancer properties.

Considering the cytotoxical effects of different Eryngium species, it seems reasonable to evaluate the cytotoxic activity of E. thyrsoideum fractions on the BC, including MCF-7 and MDA-MB-231 cell lines.

The shoots of E. thyrsoideum were extracted by Soxhlet apparatus with n-Hexane, methanol, and dichloromethane solvents. Methanolic extract (strong extract) on C18 Sep-Pak column with a mobile phase of methanol-water was fractionated. Then, the cytotoxicity of different fractions of the strong extract against normal and BC cells was evaluated for 24 and 48 hours using the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay. Induction of apoptosis was assessed by flow cytometry using staining of cells treated with Annexin V/PI.

The 80% fraction of the methanolic extract illustrated the strongest cytotoxic effectiveness among the others. This strong fraction specifically prohibited the MCF-7 and MDA-MB-231 growth with minimal effect on normal cells. The prohibition of cell growth had a time- and dose-dependent manner (P < 0.001). In addition, the flow cytometric analysis indicated that the strong fraction exerted its cytotoxic effects by inducing apoptosis on the cancer cell line.

According to our results, due to effective secondary metabolites, 80% fraction of methanolic extract prohibited the growth of both types of BC cells by inducing apoptosis and had less toxicity on normal cells.

Fibromodulin (FMOD) is a secretory protein which is considered a major component of extracellular matrix. Its dysregulation in different types of cancer implies it as a promising target for cancer therapy. Within the scope of its rather wide expression in different tumors, we studied the expression of FMOD and the effect of anti-FMOD antibody in bladder cancer cells in order to identify new target for diagnostic and therapeutic interven-tions. We report here for the first time the expression of FMOD in bladder cancer cell lines in comparison to the normal cell line and tissues.

A peptide-based produced anti-FMOD murine monoclonal antibody (mAb) (clone 2C2-A1) was applied for evaluation of FMOD expression in bladder cancer and normal tissues by immunohistochemistry (IHC) staining. Furthermore, the expression of FMOD was examined in human bladder cell lines, 5637 and EJ138, as well as a non-cancerous human cell line, human fetal foreskin fibroblast (HFFF), by immunocytochemistry (ICC) and flow cytometry. The apoptosis induction of anti-FMOD mAb was also evaluated in bladder cancer cells.

IHC and ICC analyses revealed that the qualitative expression of FMOD in bladder cancer tissues and cell lines is higher than in normal tissues and cell lines. Flow cytometry analyses revealed that 2C2-A1 mAb could recognize FMOD expression in 84.05 ± 1.85%, 46.1 ± .4% , and 2.56 ± 1.26% of 5637, EJ138, and HFFF cells, respectively. An effective apoptosis induction was detected in 5637 and EJ138 cells with no significant effect on HFFF cell.

To our knowledge, this is for the first time reporting surface expression of FMOD in bladder cancer. This significant surface expression of FMOD in bladder cancer with no expression in normal bladder tissues and the capacity of inducing apoptosis through directed targeting of FMOD with specific monoclonal antibody might candidates FMOD as a diagnostic marker as well as a potential immunotargeting with monoclonal antibody.