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Conventional suspension cultures have been used in Acute Myeloid Leukemia (AML) research to study its biology as well as to screen any drug molecules, since its inception. Co-culture models of AML cells and other stromal cells as well as 3 dimensional (3D) culture models have gained much attention recently. These culture models try to recapitulate the tumour microenvironment and are found to be more suitable than suspension cultures. Though animal models are being used, they require more time, effort and facilities and hence, it is essential to develop cell culture models for high-throughput screening of drugs. Here, we discuss a new co-culture model developed by our research group involving acute myeloid leukemia (AML) cells and stimulated macrophages. Other studies on co-culture systems and relevance of 3D culture in leukemic research in understanding the pathology and treatment of leukemia are also reviewed.

We present the effective synthesis and structural characterization of three novel imidazolium-thiohydantoin ligands (IMTHs, 5a-c) and their Mn(iii) complexes (Mn(iii)IMTHs, 6a-c) in this study. The findings of elemental analyses, spectral analyses and magnetic measurements will be used to infer the stoichiometry, coordination styles, and geometrical aspects of Mn(iii)IMTHs. The new compounds were evaluated for their chemotherapeutic potential against ESKAPE pathogens and liver cancer (HepG2). According to the MIC and MBC values, the bactericidal and bacteriostatic activities of IMTHs have been significantly improved following coordination with the Mn(iii) ion. The MTT assay results showed that all Mn(iii)IMTHs had the potential to reduce the viability of liver carcinoma (HepG2) cells in a dose-dependent manner, with the BF

Background Hepatocellular carcinoma (HCC) is known to be the second leading cause of cancer-related mortality worldwide. For improving the prognosis as well as reducing the rate of mortality, early diagnosis of HCC is a must. Aims This study was conducted to assess the ability of the serum expression of exosomal miR-18a and miR-222 to differentiate and diagnose patients with HCC, patients with liver cirrhosis, and healthy controls. Methods This study included 51 patients with liver cirrhosis, 51 patients with HCC on top of hepatitis C virus (HCV) infection, and 50 healthy controls. Results miR-18a and miR-222 were assessed using reverse transcription-polymerase chain reaction. MiR-18a and miR-222 levels were significantly higher in the liver cirrhosis and HCC groups than the control group ( p ˂ 0.001). However, no statistically significant difference was found between patients with HCC and liver cirrhosis ( p = 0.4 for miR-18a and p = 0.1 for miR-222). ROC curve analyses to evaluate the diagnostic performances of the two miRNAs as important noninvasive diagnostic markers revealed a best cutoff value of 2 for miR-18a to differentiate between liver cirrhosis, HCC, and healthy controls. And for mir-222, a cutoff value of 1.7 and 1.9 showed the highest specificity for discrimination between liver cirrhosis, HCC, and healthy controls, respectively. Moreover, logistic regression model revealed that miR-18a expression was independent predictive factor in HCC patients ( p = 0.004), while miR-222 expression was independent predictive factor in liver cirrhosis patients ( p < 0.001). Conclusion miR-18a and miR-222 were significantly discriminative markers between patients with liver cirrhosis and HCC and healthy individuals. Therefore, they have a prognostic rather than a diagnostic value. Moreover, miR-18a and miR-222 could be useful in identifying liver injuries, including fibrosis and cirrhosis.

The present work studied the antibacterial effect of different formulations of Chitosan (Chs); free Thymoquinone (Tq), and Thymoquinone-loaded chitosan nanoparticles (Tq-Chs NPs) on Bombyx mori L. infected by pathogenic bacteria, by evaluating some biochemical aspects of larval haemolymph. Bombyx mori larvae classified into three groups; each group was fed on mulberry leaves treated with one of the formulations (free Tq, free Chs and Tq-encapsulated Chs) with drug concentrations (0.05% and 0.1%). Before being used, the prepared formulations were characterized by size, morphology, encapsulation efficiency; in addition, in vitro drug release experiment was performed. The biochemical results revealed an improvement in the antibacterial and antioxidant activity of the studied formulations, especially with increasing the concentrations compared to control; (Tq-encapsulated Chs) followed by free Chs and free Tq revealed highly significant decreases in mixed function oxidase (MFO) activity, especially larvae treated with the high concentration (0.1%). The activity of acid phosphatase, total lipid content and carbohydrates pointed to a highly significant increase. Similarly, the qualitative analysis of silkworm proteins showed an obvious variation in the number and position of the detected protein bands from one treatment to another. It can be concluded that Tq and Chs showed an inhibitory impact against pathogenic bacteria infecting Bombyx mori larvae, proved their effects as antioxidant and anti-inflammatory agents which could be improved by loading Tq on Chs nanoparticles (Tq-Chs NPs).

CD47 is over-expressed in Acute Myeloid Leukemia (AML) and functions as an inhibitory signal, suppressing phagocytosis by binding to signal regulatory protein α (SIRPα) on the surface of macrophages. Inhibition of CD47 restores the immune surveillance of AML cells. However, the inhibition of CD47 in AML by activated macrophages and the subsequent effects on different immune response parameters are not fully understood. Here, we demonstrate the use of a distinct co-culture method to inhibit CD47 and therefore eliminate AML cells by macrophages in vitro. Human chemically induced THP-1 macrophages were activated using different concentrations of lipopolysaccharide (LPS) and co-culturing with three AML cancer cell lines (HL-60, NB4, and THP-1), respectively, as well as normal human peripheral blood mononuclear cells (PBMC). CD47 inhibition was observed in and selective to AML but not observed in normal PBMC. Additionally, calreticulin (CRT) levels were elevated in the same cell lines simultaneously, after co-culturing with activated human macrophages, but not elevated in normal cells. We also show that the activated macrophages secreted high levels of cytokines, including IL-12p70, IL-6, and TNF-α, consistent with the elimination of AML by macrophages. Our study reveals the potential of this model for screening new drugs against AML and the possibility of using human macrophages in AML treatment in the future.

Background The association between androgenetic alopecia (AGA) and cardiovascular risk is still controversial. Objective To evaluate the association between AGA and the risk of cardiovascular diseases (CVDs) using lipoprotein a [Lp (a)] and high-sensitivity C-reactive protein (hs-CRP) as biomarkers of cardiovascular risk. Patients and methods A case–control study was done on 30 patients with AGA and 25 healthy controls with normal hair status. Both patients and their controls were examined generally and locally. Classification of AGA was done according to Hamilton–Norwood scale for male patients and Ludwig’s classification for women. Hair examination was confirmed using dermoscopy for the assessment of signs of AGA. All participants were investigated for serum lipids including cholesterol, triglycerides, low-density lipoprotein, high-density lipoprotein, and Lp (a), in addition to hs-CRP. Results After exclusion of the well-known risk factors for CVD, AGA patients showed higher positive family history of CVD in first-degree relatives than the controls. Serum lipids including serum Lp (a) and hs-CRP were significantly higher in AGA patients. Lp (a) was positively correlated with hs-CRP in AGA patients. Both Lp (a) and hs-CRP were positively correlated with the severity of AGA in both men and women. Both markers did not correlate significantly with lipid parameters including cholesterol, triglycerides, low-density lipoprotein, and high-density lipoprotein. There was no significant difference between men and women with AGA as regards the studied parameters. Both hs-CRP and Lp (a) showed significant discrimination using area under the receiver operating characteristic curve where Lp (a) showed area under the curve=0.928 with 95% confidence interval, 0.863–0.993 and P=0.001 while hs-CRP showed area under the curve=0.963 with 95% confidence interval 0.921–1.0, and P value less than 0.001. Multivariate logistic regression analysis showed that hs-CRP was the only significant independent factor affecting cardiac risk in AGA cases. Conclusion AGA patients showed higher risk for CVD. Both Lp (a) and hs-CRP can be used as a predictor for cardiovascular risk in such patients; however, hs-CRP is still considered an independent risk factor for cardiovascular insults in AGA patients.

Cryptosporidium spp. are one of the most important waterborne pathogens worldwide and a leading cause of mortality from waterborne gastrointestinal diseases. Detection of Cryptosporidium spp. in water can be very challenging due to their low numbers and the complexity of the water matrix. This review describes the biology of Cryptosporidium spp. and current methods used in their detection with a focus on C. parvum and C. hominis. Among the methods discussed and compared are microscopy, immunology-based methods using monoclonal antibodies, molecular methods including PCR (polymerase chain reaction)-based assays, and emerging aptamer-based methods. These methods have different capabilities and limitations, but one common challenge is the need for better sensitivity and specificity, particularly in the presence of contaminants. The application of DNA aptamers in the detection of Cryptosporidium spp. oocysts shows promise in overcoming these challenges, and there will likely be significant developments in aptamer-based sensors in the near future.

Optical fiber-based surface plasmon resonance (OF-SPR) sensors have demonstrated high versatility and performances over the last years, which propelled the technique to the heart of numerous and original biosensing concepts. In this work, we contribute to this effort and present our recent findings about the detection of breast cancer HER2 biomarkers through OF-SPR optrodes. 1 cm-long sections of 400 μm core-diameter optical fibers were covered with a sputtered gold film, yielding enhanced sensitivity to surface refractive index changes. Studying the impacts of the gold film thickness on the plasmonic spectral response, we improved the quality and reproducibility of the sensors. These achievements were correlated in two ways, using both the central wavelengths of the plasmon resonance and its influence on the bulk refractive index sensitivity. Our dataset was fed by additional biosensing experiments with a direct and indirect approach, relying on aptamers and antibodies specifically implemented in a sandwich layout. HER2 biomarkers were specifically detected at 0.6 μg/mL (5.16 nM) in label-free while the amplification with HER2-antibodies provided a nearly hundredfold signal magnification, reaching 9.3 ng/mL (77.4 pM). We believe that these results harbinger the way for their further use in biomedical samples.

In the biomedical detection context, plasmonic tilted fiber Bragg gratings (TFBGs) have been demonstrated to be a very accurate and sensitive sensing tool, especially well-adapted for biochemical detection. In this work, we have developed an aptasensor following a triple strategy to improve the overall sensing performances and robustness. Single polarization fiber (SPF) is used as biosensor substrate while the demodulation is based on tracking a peculiar feature of the lower envelope of the cladding mode resonances spectrum. This method is highly sensitive and yields wavelength shifts several tens of times higher than the ones reported so far based on the tracking of individual modes of the spectrum. An amplification of the response is further performed through a sandwich assay by the use of specific antibodies. These improvements have been achieved on a biosensor developed for the detection of the HER2 (Human Epidermal Growth Factor Receptor-2) protein, a relevant breast cancer biomarker. These advanced developments can be very interesting for point-of-care biomedical measurements in a convenient practical way.

CD47 is over-expressed in Acute Myeloid Leukemia (AML) and functions as an inhibitory signal, suppressing phagocytosis by binding to signal regulatory protein α (SIRPα) on the surface of macrophages. Inhibition of CD47 restores the immune surveillance of AML cells. However, the inhibition of CD47 in AML by activated macrophages and the subsequent effects on different immune response parameters are not fully understood. Here, we demonstrate the use of a distinct co-culture method to inhibit CD47 and therefore eliminate AML cells by macrophages in vitro. Human chemically induced THP-1 macrophages were activated using different concentrations of lipopolysaccharide (LPS) and co-culturing with three AML cancer cell lines (HL-60, NB4, and THP-1), respectively, as well as normal human peripheral blood mononuclear cells (PBMC). CD47 inhibition was observed in and selective to AML but not observed in normal PBMC. Additionally, calreticulin (CRT) levels were elevated in the same cell lines simultaneously, after co-culturing with activated human macrophages, but not elevated in normal cells. We also show that the activated macrophages secreted high levels of cytokines, including IL-12p70, IL-6, and TNF-α, consistent with the elimination of AML by macrophages. Our study reveals the potential of this model for screening new drugs against AML and the possibility of using human macrophages in AML treatment in the future.

The detection of circulating tumor cells (CTCs) represents an important goal in oncological diagnosis and treatment, as CTCs are responsible for metastasis in several forms of cancer and are present at very low concentration. Their detection should occur at around 1-10 cells/mL of blood for diagnosis purpose. In this work, we propose an all-fiber plasmonic aptasensor featuring multiple narrowband resonances in the near-infrared wavelength range to detect metastatic breast cancer cells. To this aim, specific aptamers against mammaglobin-A proteins were selected and immobilized as bioreceptors on the optical fiber surface. In vitro assays confirm that label-free and real-time detection of cancer cells (LOD of 49 cells/mL) occurs within 5 minutes, while the additional use of functionalized gold nanoparticles allows a two-fold amplification of the biosensor response. Differential measurements on selected optical resonances were used to process the sensor response and results were confirmed by microscopy analysis. The detection of only 10 cancer cells/mL was performed with relevant specificity against non-target cells with comparable sizes and shapes.

The detection of circulating tumor cells (CTCs), which are responsible for metastasis in several forms of cancer, represents an important goal in oncological diagnosis and treatment. These cells remain extremely challenging to detect, despite numerous previous studies, due to their low concentration (1-10 cells/mL of blood). In this work, an all-fiber plasmonic aptasensor featuring multiple narrowband resonances in the near-infrared wavelength range was developed to detect metastatic breast cancer cells. To this aim, specific aptamers against mammaglobin-A were selected and immobilized as receptors on the sensor surface. In vitro assays confirm that the label-free and real-time detection of cancer cells [limit of detection (LOD) of 49 cells/mL] occurs within 5 min, while the additional use of functionalized gold nanoparticles allows a 2-fold amplification of the biosensor response. Differential measurements on selected optical resonances were used to process the sensor response, and results were confirmed by microscopy. The detection of only 10 cancer cells/mL was achieved with relevant specificity against control cells and with quick response time.

AIM To investigate how Tregs are regulated in chronic hepatitis C virus (HCV) patients via assessment of Tregs markers (granzyme 2, CD69 and FoxP3), Teffs markers [TNFRSF4 (OX40), INFG] and CD4, CD25 genes. METHODS A prospective study was conducted on 120 subjects divided into 4 groups: Group I (n = 30) treatment naïve chronic HCV patients; Group II (n = 30) chronic HCV treated with Peg/Riba; Group III (n = 30) chronic HCV associated with non-organ specific autoantibody and Group IV (n = 30) healthy persons as a control group. Tregs and Teffs markers were assessed in peripheral blood mononuclear cells by quantitative real time reverse transcriptase-polymerase chain reaction. RESULTS Chronic HCV patients exhibited significant higher levels of both Teffs and Tregs in comparison to healthy control group. Tregs markers were significantly decreased in Peg/Riba treated HCV patients in comparison to treatment naïve HCV group. In HCV patients with antinuclear antibody (ANA) +ve, Tregs markers were significantly decreased in comparison to all other studied groups. Teffs markers were significantly elevated in all HCV groups in comparison to control and in HCV group with ANA +ve in comparison to treatment naïve HCV group. CONCLUSION Elevated Tregs cells in chronic HCV patients dampen both CD4+ and CD8+ autologous T cell immune response. Interferon-α and ribavirin therapy suppress proliferation of Tregs. More significant suppression of Tregs was observed in HCV patients with autoantibodies favoring pathological autoimmune response.

Tilted fiber Bragg gratings (TFBGs) are now a well-established technology in the scientific literature, bringing numerous advantages, especially for biodetection. Significant sensitivity improvements are achieved by exciting plasmon waves on their metal-coated surface. Nowadays, a large part of advances in this topic relies on new strategies aimed at providing sensitivity enhancements. In this work, TFBGs are produced in both single-mode and multimode telecommunication-grade optical fibers, and their relative performances are evaluated for refractometry and biosensing purposes. TFBGs are biofunctionalized with aptamers oriented against HER2 (Human Epidermal Growth Factor Receptor-2), a relevant protein biomarker for breast cancer diagnosis. In vitro assays confirm that the sensing performances of TFBGs in multimode fiber are higher or identical to those of their counterparts in single-mode fiber, respectively, when bulk refractometry or surface biosensing is considered. These observations are confirmed by numerical simulations. TFBGs in multimode fiber bring valuable practical assets, featuring a reduced spectral bandwidth for improved multiplexing possibilities enabling the detection of several biomarkers.

Mammaglobin B (MGB2) and mammaglobin A (MGB1) are proteins expressed in metastatic breast cancers. The early detection of circulating tumor cells (CTCs) in breast cancer patients is crucial to decrease mortality rate. Herein, novel aptamers were successfully selected and characterized against MGB2 and MGB1 proteins using a hybrid SELEX approach. The potential use of the selected aptamers in breast CTC detection was studied using spiked breast cancer cells in whole blood lysate. The results obtained from this study showed that the selected aptamers (MAMB1 and MAMA2) bind to their target breast cancer cell lines with high affinity (low nanomolar Kd values) and specificity. They also bind to their free recombinant target proteins and show minimal non-specific binding to normal and other cancer cell lines. Additionally, they were able to distinguish a low number of breast cancer cells spiked in whole blood lysate containing normal blood cells. The results obtained in this study indicate the great potential for the use of aptamers to detect MGB1 and MGB2 protein biomarkers, expressed on the surface of breast CTCs.

Background Psoriasis is a chronic inflammatory disease that poses a challenge in its pathogenesis and treatment. Calprotectin is upregulated in psoriatic epidermis and is supposed to have a role in the inflammatory process of psoriasis. Calprotectin was suggested as a biomarker to detect responders to methotrexate in juvenile idiopathic arthritis. Objective To evaluate the relation of calprotectin serum level and psoriasis severity and to assess if it can predict response to methotrexate and disease relapse after stoppage of the treatment. Patients and methods Thirty patients with psoriasis vulgaris and 30 controls were included. Serum calprotectin was measured in controls and in patients before and after methotrexate therapy for 3 months. The patients were followed up for 6 months to detect disease stability or relapse and its relation to calprotectin level at the end of therapy. Results Higher calprotectin level in psoriatic patients was detected before methotrexate treatment than after treatment. Significant positive correlation was found between serum calprotectin and psoriasis area and severity index score. Calprotectin was higher in patients before starting methotrexate in those defined as responders than nonresponders (P=0.015) and can predict response to treatment with a cutoff value of 60 ng/ml (sensitivity 82.35% and specificity 69.23%). The level of calprotectin was higher in patients during their clinical remission who relapsed after stoppage of methotrexate than the nonrelapsed cases (P=0.021) and can predict the relapse with a cutoff value of 56.5 ng/ml (sensitivity 66.67% and specificity 100%). Conclusion Calprotectin can be a marker of psoriasis severity. Higher level of baseline serum calprotectin can predict psoriatic patients who will improve on methotrexate therapy. In addition, higher calprotectin levels were associated with relapse risk after stoppage of methotrexate.

Introduction Diabetes mellitus (DM) is a multiorgan disease that leads to neurovascular complications that disturb the normal erectile function. Aim The aim of the current work was to study the early changes occurring in the level of calcitonin gene-related peptide (CGRP) and histopathological changes in penile tissues of uncontrolled diabetic rats. Materials and Methods This study was carried on 50 adult male Sprague-Dawley rats divided into two main groups: group I (control, n = 10) and group II (diabetic, n = 40). Type I DM was induced by a single intraperitoneal injection of streptozotocin (60 mg/kg). The tissue level of CGRP and histopathological examination of rat penises were assessed at 2, 4, 6, and 8 weeks after induction of DM. Results CGRP was higher in the diabetic group at 4, 6, and 8 weeks than in the control group. However, endothelial changes and decreased smooth muscles mass started only 2 weeks after induction of DM. Conclusion Deterioration of histopathological features of the uncontrolled diabetic rats corporeal tissues is time dependent. Furthermore, vascular changes seem to precede the neurological changes.

DNA aptamers were developed against murine norovirus (MNV) using SELEX (Systematic Evolution of Ligands by EXponential enrichment). Nine rounds of SELEX led to the discovery of AG3, a promising aptamer with very high affinity for MNV as well as for lab-synthesized capsids of a common human norovirus (HuNoV) outbreak strain (GII.3). Using fluorescence anisotropy, AG3 was found to bind with MNV with affinity in the low picomolar range. The aptamer could cross-react with HuNoV though it was selected against MNV. As compared to a non-specific DNA control sequence, the norovirus-binding affinity of AG3 was about a million-fold higher. In further tests, the aptamer also showed nearly a million-fold higher affinity for the noroviruses than for the feline calicivirus (FCV), a virus similar in size and structure to noroviruses. AG3 was incorporated into a simple electrochemical sensor using a gold nanoparticle-modified screen-printed carbon electrode (GNPs-SPCE). The aptasensor could detect MNV with a limit of detection of approximately 180 virus particles, for possible on-site applications. The lead aptamer candidate and the aptasensor platform show promise for the rapid detection and identification of noroviruses in environmental and clinical samples.

Background Thrombolytic therapy is currently the only FDA-approved treatment for acute ischemic stroke. Hence, early diagnosis and risk stratification is of great importance in management. Objective The aim of this work was to study serum level of matrix metalloproteinase-9 (MMP-9) within 24 h of acute ischemic stroke onset and its relation with clinical severity. Patients and methods Thirty patients with acute ischemic stroke were subjected to measurement of serum MMP-9 within 24 h of stroke onset and clinical assessment of stroke severity. Thirty healthy volunteers of matched age and sex were included as controls. Results Fifteen male and 15 female patients with a mean age of 61 ± 7.11 years were studied. The mean National Institutes of Health Stroke Scale (NIHSS) score on admission was 11.17 ± 4.76. The mean serum level of MMP-9 in patients was 998.8 ± 154.72 ng/ml, which was significantly higher compared with the serum level of MMP-9 in controls ( P = 0.003). The mean NIHSS of patients with normal serum level of MMP-9 was less than the mean NIHSS in patients with high MMP-9 serum levels ( P = 0.003). There was a significant positive correlation between serum level of MMP-9 and NIHSS score ( r = 0.5; P = 0.005) even after adjustment of other variables )age, sex, diabetes, hypertension, fasting blood sugar, uric acid, serum triglycerides, serum cholesterol, and right and left carotid intima media thickness( ( r = 0.48; P = 0.032). Conclusion Serum MMP-9 level was found to be high in acute ischemic stroke patients and correlated with clinical stroke severity.