Heterotypic tumor spheroids: a platform for nanomedicine evaluation.

Nanomedicine has emerged as a promising therapeutic approach, but its translation to the clinic has been hindered by the lack of cellular models to anticipate how tumor cells will respond to therapy. Three-dimensional (3D) cell culture models are thought to more accurately recapitulate key features of primary tumors than two-dimensional (2D) cultures. Heterotypic 3D tumor spheroids, composed of multiple cell types, have become more popular than homotypic spheroids, which consist of a single cell type, as a superior model for mimicking in vivo tumor heterogeneity and physiology. The stromal interactions demonstrated in heterotypic 3D tumor spheroids can affect various aspects, including response to therapy, cancer progression, nanomedicine penetration, and drug resistance. Accordingly, to design more effective anticancer nanomedicinal therapeutics, not only tumor cells but also stromal cells (e.g., fibroblasts and immune cells) should be considered to create a more physiologically relevant in vivo microenvironment. This review aims to demonstrate current knowledge of heterotypic 3D tumor spheroids in cancer research, to illustrate current advances in utilizing these tumor models as a novel and versatile platform for in vitro evaluation of nanomedicine-based therapeutics in cancer research, and to discuss challenges, guidelines, and future directions in this field.

An agarose-alginate microfluidic device for the study of spheroid invasion, ATRA inhibits CAFs-mediated matrix remodeling.

Growing evidence demonstrates that cancer-associated fibroblasts (CAF) are responsible for tumor genesis, growth, metastasis, and treatment response. Therefore, targeting these cells may contribute to tumor control. It has been proposed that targeting key molecules and pathways of proliferative functions can be more effective than killing CAFs. In this regard, multicellular aggregates, like spheroids, can be used as human tumor models. Spheroids closely resemble human tumors and mimic many of their features. Microfluidic systems are ideal for cultivation and study of spheroids. These systems can be designed with different biological and synthetic matrices in order to have a more realistic simulation of the tumor microenvironment (TME). In this study, we investigated the effect of all-trans retinoic acid (ATRA) on 3D spheroid invasion of MDA-MB cells exposed to hydrogel matrix derived from CAFs. The number of invasive cells significantly decreased in CAF-ECM hydrogel treated with ATRA (p < 0.05), which indicates that ATRA could be effective for CAFs normalization. This experiment was done using an agarose-alginate microfluidic chip. As compared with common methods, such hydrogel casting is an easier method for chip fabrication and can even reduce costs. Supplementary Information: The online version contains supplementary material available at 10.1007/s10616-023-00578-y.

Middle Ear Status in Cleft Lip and Palate Patients: A Five-Year Follow-Up.

Introduction: The best strategy to treat otitis media with effusion in cleft lip/palate patients is still under debate. This research aimed to evaluate the otologic outcomes in children at least five years post-repair. Materials and Methods: A retrospective study was conducted on 40 children who underwent palatoplasty between January 1, 2012, and January 1, 2014, at Children's Medical Center (Tehran, Iran). Patients had intervelar veloplasty under magnification (Sommerlad's Technique). Based on patients' charts, their age, gender, cleft type, date of palatoplasty, as well as the date and the frequency of ventilation tube (VT) insertion, were recorded. Furthermore, otomicroscopy, middle ear status, and tympanometry were assessed five years postoperatively. Results: There was no significant difference in middle ear status between children with complete and incomplete cleft palates. The mean age at the time of study and the mean follow-up duration were significantly higher in the normal middle ear group, compared to the abnormal middle ear group (7.7±1.6 vs. 6.8±0.9, P=0.03 and 6±1.15 vs. 5.42±0.9, P=0.04, respectively). Middle ear status was not significantly different between early or late palatoplasty patients. In addition, the frequency and timing of VT insertion were not significantly different between the two groups. Conclusions: Middle ear status improved as patients grew older; however, the age of palatoplasty and the frequency of VT insertion were not significant prognostic factors in patients who underwent intervelar veloplasty under magnification.

Antiproliferative activity of new derivatives of pyrazino[1,2-a]benzimidazole: Integrated cell-based assay and computational studies with divalent magnesium, iron, and copper ions.

Magnesium, iron, and copper are three vital metals that play essential roles in cancer cell proliferation. This study aimed to evaluate the metal chelation of new derivatives of pyrazino[1,2-a]benzimidazole and investigate their antiproliferative properties. The density functional theory method has been employed to evaluate the complexation properties of new synthetic pyrazino[1,2-a]benzimidazole derivatives possessing the 4-OMe, 2,4-dimethyl, and 3,4,5-trimethoxy substitution on N-2 phenyl ring with divalent magnesium, iron, and copper. The free energies for the water-ligand exchange reactions were employed to investigate the thermodynamic stability, water exchange properties, and electronic properties in the gas phase. Natural population analysis was employed to estimate atomic partial charges, second-order interactions between the filled and vacant orbitals, and the occupancies of the metals' valence s, p, and d orbitals. Among pyrazino[1,2-a]benzimidazole derivatives, the 3,4,5-trimethoxy substituted pyrazino[1,2-a]benzimidazole shows better electron donor ability. This compound also reduced proliferation and increased the apoptosis of human glioblastoma cancer cells.

An insight into the potentials of carbon dots for in vitro live-cell imaging: recent progress, challenges, and prospects.

Carbon dots (CDs) are a strong alternative to conventional fluorescent probes for cell imaging due to their brightness, photostability, tunable fluorescence emission, low toxicity, inexpensive preparation, and chemical diversity. Improving the targeting efficiency by modulation of the surface functional groups and understanding the mechanisms of targeted imaging are the most challenging issues in cell imaging by CDs. Firstly, we briefly discuss important features of fluorescent CDs for live-cell imaging application in this review. Then, the newest modulated CDs for targeted live-cell imaging of whole-cell, cell organelles, pH, ions, small molecules, and proteins are elaborately discussed, and their challenges in these fields are explained.

Investigation of anti-cancer effects of new pyrazino[1,2-a]benzimidazole derivatives on human glioblastoma cells through 2D in vitro model and 3D-printed microfluidic device.

AIMS: Recent studies show targeted therapy of new pyrazino[1,2-a]benzimidazole derivatives with COX-II inhibitory effects on different cancer cells. This study aimed to investigate 2D cell culture and 3D spheroid formation of glioblastoma multiforme (GBM) cells using a microfluidic device after exposure to these compounds. MAIN METHODS: After isolating astrocytes from human GBM samples, IC50 of 2,6-dimethyl pyrazino[1,2-a]benzimidazole (L1) and 3,4,5-trimethoxy pyrazino[1,2-a]benzimidazole (L2) were determined as 13 µM and 85 µM, respectively. Then, in all experiments, cells were exposed to subtoxic concentrations of L1 (6.5 µM) and L2 (42.5 µM), which were ½IC50. In the following, in two phases, cell cycle, migration, and gene expression through 2D cell culture and tumor spheroid formation ability using a 3D-printed microfluidic chip were assessed. KEY FINDINGS: The obtained results showed that both compounds have positive effects in reducing G2/M cell population and GBM cell migration. Furthermore, real-time gene expression data showed that L1 and L2 significantly impact the upregulation of P21 and P53 and down-regulation of cyclin D1, MMP2, and MMP9. On the other hand, GBM spheroids exposed to L1 and L2 become smaller with fewer live cells. SIGNIFICANCE: Our data on human isolated astrocyte cells in 2D and 3D cell culture conditions showed that L1 and L2 compounds could reduce GBM cells' invasion by controlling gene expressions associated with migration and proliferation. Moreover, designing microfluidic platform and related cell culture protocols facilitates the broad screening of 3D multicellular tumor spheroids derived from GBM tumor biopsies and provides effective drug development for brain gliomas.

The Preparedness of Primary Health Care Network in terms of Emergency Risk Communication: A Study in Iran.

OBJECTIVE: Emergency Risk Communication (ERC) is known as 1 of the important components of an effective response to public health emergencies. In this study, we aimed to investigate the preparedness of the Primary Health Care Network (PHCN) of Iran in terms of the ERC. METHODS: This study was conducted in 136 Primary Health Care Facilities (PHCFs) affilated to Shahrekord University of Medical Sciences, Chaharmahal and Bakhtiari Province, Iran. Data in terms of ERC were collected using a checklist developed by the Center of Disease Control and Prevention (CDC). RESULTS: The findings of the study revealed that 65.9% of the PHCFs had low preparedness in terms of the ERC, 33.3% had a moderate level and 0.8% had high preparedness in this regard. There was a significant difference between the level of ERC and the history of crisis in the past year, PHCF type, and the education level of the responsible employees in the crisis unit in the PHCF. CONCLUSIONS: The results showed that the PHCFs studied need to increase their capacity and capability in the field of ERC. Further efforts to provide ERC components may increase the preparedness of PHCN in Iran in terms of the ERC.

Assessment of cytotoxic effects of new derivatives of pyrazino[1,2-a] benzimidazole on isolated human glioblastoma cells and mitochondria.

AIMS: Glioblastoma multiforme (GBM) is a highly devastating malignant brain tumor with poor pharmacotherapy. Based on COX-2 inhibitory effects in preventing cancer progression, new pyrazino[1,2-a]benzimidazole derivatives were assessed on isolated human GBM cells. MAIN METHODS: In this study, firstly, primary culture of astrocytes from human GBM samples was prepared and exposed to 2,6-dimethyl pyrazino[1,2-a]benzimidazole (L1) and 3,4,5-trimethoxy pyrazino[1,2-a]benzimidazole (L2) for finding their half-maximal inhibitory concentration (IC50). In the following, in two phases, cell apoptosis pathway and mitochondrial markers were investigated on GBM and also HEK293 cells (as non-cancerous normal cells). KEY FINDINGS: The MTT results represented a remarkable selective cytotoxic effect of both L1 and L2 on GBM cells, and interestingly not on normal cells. After 48 h, IC50 of L1 and L2 were calculated as 13 µM and 85 µM, respectively. Annexin/PI staining showed that L1 and L2 induce apoptosis in GBM cells, and caspase measurement showed that apoptosis occurs through mitochondrial signaling. In the clonogenic assay, GBM cells formed more paraclones and fewer holoclones after treating with L1 and L2. L1 and L2 also selectively enhanced mitochondrial damaged markers, including reactive oxygen species (ROS) formation, and mitochondrial swelling, decreased mitochondrial membrane potential (MMP) and cytochrome c release in isolated cancerous GBM mitochondria. SIGNIFICANCE: Our findings on human primary astrocyte cells illustrated that L1 and L2 compounds, with COX-2 inhibitory effect, through the intrinsic pathway of apoptosis concerning mitochondrial damage enhancement have therapeutic potentials on GBM.

A combined 3D printing/CNC micro-milling method to fabricate a large-scale microfluidic device with the small size 3D architectures: an application for tumor spheroid production.

The fabrication of a large-scale microfluidic mold with 3D microstructures for manufacturing of the conical microwell chip using a combined projection micro-stereolithography (PµSL) 3D printing/CNC micro-milling method for tumor spheroid formation is presented. The PµSL technique is known as the most promising method of manufacturing microfluidic chips due to the possibility of creating complex three-dimensional microstructures with high resolution in the range of several micrometers. The purpose of applying the proposed method is to investigate the influence of microwell depths on the formation of tumor spheroids. In the conventional methods, the construction of three-dimensional microstructures and multi-height chips is difficult, time-consuming, and is performed using a multi-step lithography process. Microwell depth is an essential parameter for microwell design since it directly affects the shear stress of the fluid flow and the diffusion of nutrients, respiratory gases, and growth factors. In this study, a chip was made with microwells of different depth varying from 100 to 500 µm. The mold of the microwell section is printed by the lab-made PµSL printer with 6 and 1 µm lateral and vertical resolutions. Other parts of the mold, such as the main chamber and micro-channels, were manufactured using the CNC micro-milling method. Finally, different parts of the master mold were assembled and used for PDMS casting. The proposed technique drastically simplifies the fabrication and rapid prototyping of large-scale microfluidic devices with high-resolution microstructures by combining 3D printing with the CNC micro-milling method.

The Association Between Demographic Characteristics and Attempting of Pregnancy with Postpartum Depression and Anxiety Among Women Referring to Community Health Centres: A Cross Sectional Study.

BACKGROUND: Postpartum depression (PPD) and anxiety are considered as a risk factor for mother and infant health. Therefore, the present study aims to explore the association between demographic characteristics and pregnancies with PPD and anxiety. METHODS: A cross-sectional study was conducted on 400 Iranian women referring to health centres of the Zarand City four weeks to six months from the date of their childbirth, in the first half of 2018. RESULT: The results showed that employed women with pregnancies who were categorised as depression and anxiety were more likely to have low gestational age, food insecurity, several deliveries, cesarean delivery and unintended pregnancy as well as they were not satisfied with their infant's gender. Also, women with several deliveries had lower risk for PPD before and after adjustment for confounders (odds ratio [OR] = 0.92, 95% confidence interval [CI]: 0.88-0.97, P < 0.001) and had lower risk for postpartum anxiety only after adjustment for confounders (OR = 0.82, 95% CI: 0.75-0.89, P < 0.001). CONCLUSION: Eventually, demographic characteristics and attempting of pregnancy were independently associated with PPD and postpartum anxiety in women. There need to be more social and governmental support of employed women after delivery to decrease their occupational stresses to deal with PPD and anxiety in the studied population.

Carbon dots with pH-responsive fluorescence: a review on synthesis and cell biological applications.

This review summarizes state of the art synthesis and applications of carbon dots (CDs) with pH-responsive fluorescence. Following an introduction, the first section covers methods for the preparation of pH-responsive CDs, with subsections on general methods for preparing CDs (by hydrothermal, solvothermal, electrochemical, microwave, laser ablation, pyrolysis or chemical oxidation polymerization methods), and on precursors for synthesis. This is followed by a section on the mechanisms of pH-responsivity (by creating new functional groups, change of energy levels, protonation and deprotonation, aggregation, or by introduction shells). Several Tables are presented that give an overview of the wealth of methods and materials. A final section covers applications of carbon dots (CDs) with pH-responsive fluorescence for sensing, drug delivery, and imaging. The conclusion summarizes the current status, addresses challenges, and gives an outlook on potential future trends. Graphical abstract The synthesis and biological applications of carbon dots(CDs) with pH-responsive fluorescence are summarized. Precursors and methods for preparation of pH-responsive CDs, mechanisms of pH-responsivity, and biological applications of CDs with pH-responsive fluorescence for sensing, drug delivery, and imaging are discussed.

Effect of carbonization degree of carbon dots on cytotoxicity and photo-induced toxicity to cells.

BACKGROUND: Pristine carbon dots (CDs) derived from citric acid pyrolysis are used in a variety of biomedical research such as imaging and drug delivery. However, potential cytotoxic effects of pyrolysis temperature on cells is underexplored. To address this need, we studied toxicity of the CDs to breast cancer cells using MTT and LDH assays. In addition, we investigated photo-induced cytotoxicity of the synthesized CDs in a wide concentration range under white light. RESULTS: Our results suggest little cytotoxicity of the CDs after 24 h exposure of cells. Only the high quantum yield CDs caused a significant toxicity to cells at the highest concentrations of 2.0 and 1.5 mg/ml compared to other CDs at similar concentrations. The synthesized CDs entered the cells without any significant cytotoxicity. The CDs also caused a concentration- and irradiation time-dependent photo-induced cytotoxicity. CONCLUSION: The optimization of synthesis conditions from this study may help develop safe and efficient CDs for imaging and drug delivery.

Physicochemical and cytotoxicity analysis of green synthesis carbon dots for cell imaging.

Carbon dots (CDs) have outstanding optical properties, biocompatibility, and photostability, making them attractive for imaging applications. A facile and green one-step hydrothermal synthesis method is proposed, which can be safely used in a wide range of applications such as chemical sensing, bioimaging, and optoelectronics. In this study, we report green synthesis of carbon dots from bitter orange juice (Citrus Aurantium) by hydrothermal treatment for the first time. We studied effects of time, temperature, and pH on fluorescence of CDs, characterized them using various spectroscopic and microscopic methods, and evaluated their toxicity to different cell lines. Identifying an optimum reaction condition of 180 ºC for 7 h heating gave CDs that showed pH-dependent fluorescence, with the largest fluorescence at a pH of 7.0. The CDs were 1-2 nm in size with a spherical morphology and negative surface charge. The CDs showed a high quantum yield of 19.9 %, reasonable photostability, excellent water solubility, and long fluorescence lifetime. A one step hydrothermal rout led to various hydrophilic functional groups on the surface of the CDs. Our results showed that the CDs were non-toxic over a large concentration range and effective for imaging of cells, indicating their potential as imaging probes in medical diagnostics and biosensor applications.

Optical Imaging Approaches to Monitor Static and Dynamic Cell-on-Chip Platforms: A Tutorial Review.

Miniaturized laboratories on chip platforms play an important role in handling life sciences studies. The platforms may contain static or dynamic biological cells. Examples are a fixed medium of an organ-on-a-chip and individual cells moving in a microfluidic channel, respectively. Due to feasibility of control or investigation and ethical implications of live targets, both static and dynamic cell-on-chip platforms promise various applications in biology. To extract necessary information from the experiments, the demand for direct monitoring is rapidly increasing. Among different microscopy methods, optical imaging is a straightforward choice. Considering light interaction with biological agents, imaging signals may be generated as a result of scattering or emission effects from a sample. Thus, optical imaging techniques could be categorized into scattering-based and emission-based techniques. In this review, various optical imaging approaches used in monitoring static and dynamic platforms are introduced along with their optical systems, advantages, challenges, and applications. This review may help biologists to find a suitable imaging technique for different cell-on-chip studies and might also be useful for the people who are going to develop optical imaging systems in life sciences studies.

On-chip analysis of carbon dots effect on yeast replicative lifespan.

Carbon dots (CDs) are promising nanomaterials for biosensing, bioimaging, and drug delivery due to their large surface area, excellent optical properties, and thermal and chemical stability. However, biosafety of CDs is still understudied, and there is not a generally accepted standard to evaluate the toxicity of CDs. We present a gradient network generator microfluidic device for dose-dependent testing of toxicity of CDs to a unicellular eukaryotic model organism, yeast Pichia pastoris. We fully characterized the microfluidic model and compare its performance with a conventional method. The gradient generator increased the contact area between the mixing species and enabled a high-throughput testing of CDs in a wide range of concentrations in cell chambers. Real time monitoring of yeast cell proliferation in the presence of CDs showed dose-dependent growth inhibition and various budding cell shape profiles. Comparing the result of microfluidic platform and conventional method revealed statistically significant differences in the proliferation rate of the cells between the two platforms. To understand the toxicity mechanism, we studied binding of CDs to P. pastoris and found increasing interactions of CDs with the cell surface at CDs larger concentrations. This study demonstrated the utility of the gradient generator microfluidic device as a convenient tool for toxicity assessment of nanomaterials at a cellular level.

Investigation the cytotoxicity and photo-induced toxicity of carbon dot on yeast cell.

Carbon dots (CDs) as a new fluorescent material with excellent water solubility, chemical inertness, and easy surface modification are a good candidate for bioimaging and biosensing due to their low toxicity and good biocompatibility. Although carbon is not an intrinsically toxic substance, carbon nanomaterials such as CDs may cause risks to human health and the potentially hazardous effects of CDs on various living systems must be completely determined. So far, cytotoxicity studies of CDs have focused on human cells and are mainly conducted on limited cell lines. In the present study, toxicity assessment of CDs was evaluated on yeast cells Pichia pastoris as a unicellular eukaryotic model. Results revealed dose-dependent toxicity of CDs on yeast cells and less relative cell growth in 25 mg/ml of CDs as compared to the control group. CDs binding curve confirmed the interaction between CDs and surface of yeast cells. SEM images showed that the CDs caused cell shrinkage and hole formation on the surface of yeast cells and also induced slightly cell deformation. It was demonstrated that CDs could generate the ROS dose-dependently. Finally, results showed the growth inhibition and ROS generation effects of CDs were enhanced at light exposure, as an important environmental factor. These findings could have important implications for applications of CDs.

New Insight into the Concept of Carbonization Degree in Synthesis of Carbon Dots to Achieve Facile Smartphone Based Sensing Platform.

Direct pyrolysis of citric acid (CA) has been proved to be a facile bottom-up technique for making pristine carbon dots (CD) with homogenous size distribution. However, limited reports are available on systematic optimization of carbonization degree. In this investigation, pyrolysis temperatures between 160 °C and 220 °C were studied, based on CA thermal decomposition path, using various heating durations. The effect of the formation of more carbonized carbon particles (MCCPs), as the major byproduct of this method, on photoluminescence properties of CDs was also considered. The NaOH amount that neutralizes the solution and the effect of dilution on the emission intensity, were introduced as simple and accessible factors for monitoring carbonization degree, and an estimate of MCCP/CD ratio, respectively. The results show that the CDs fabricated at 160 °C, 50 minutes attain almost twice higher quantum yield (QY) of 29% than highest QY reported based on pyrolysis of CA. The so-prepared CDs can be employed as excellent candidates for turn-off sensing. As a proof of concept, detection limit of 50 nM for Hg2+ was achieved using a facile and inexpensive smartphone set-up that is able to quantify and compare fluorescent intensity in several samples simultaneously.

Spectral properties and thermal stability of AS1411 G-quadruplex.

G-quadruplexes are supramolecular structures of G-rich nucleic acid, formed by non-canonical base pairing in the presence of specific environmental inducers. These structures have been vastly considered in diagnostic and therapeutic applications. However, detailed information on structure, optical properties and thermal stability of G-quadruplex potent oligonucleotides is scarce. Herein, optical properties and thermodynamic stability of AS1411 quadruplex is reported for various concentrations of potassium and lead ions. Circular dichroism showed that AS1411 ss-DNA folds into parallel conformation in the presence of metal ions and molecular crowding condition. UV-vis spectroscopy indicated formation of quadruplex and fluorescent spectroscopy revealed intercalation of PicoGreen in its structure, with enhancement of emission intensity upon increment of metal ion concentration. This investigation also proposes high-throughput and reliable analysis of AS1411 quadruplex's thermal stability by real-time PCR technique, which can be further applied for other quadruplex structures.

Content analysis of articles published in Iranian scientific nursing journals from 2009 through 2011.

BACKGROUND: Little is known about the features of Iranian nursing journals, specifically the subject areas used in articles, study designs, sampling methods, international collaboration of Iranian nursing scholars, specialty and academic rank of authors, and the most frequently contributing academic institutions in articles. OBJECTIVES: The aim of this study was to analyze the content of the articles published in Iranian scientific nursing journals. MATERIALS AND METHODS: Quantitative content analysis was implemented to study Iranian nursing journals, which were approved by the commission for accreditation and improvement of Iranian medical journals in 2011. Thus, 763 articles from six journals, published from 2009 through 2011, were investigated. Data were extracted from the abstracts and when necessary, from the full-text of articles by visiting the websites of these journals. Descriptive statistics were used to analyze the data. RESULTS: The main subjects of published articles in Iranian scientific nursing journals were consecutively renal dialysis (n = 21), intensive care unit (n = 16), nursing education (n = 15), patient satisfaction (n = 13), quality of life (n = 12), health education (n = 11), patient education (n = 11), pain (n = 10), and education (n = 9). The majority of authors had nursing and midwifery specialty (52.59%) followed by epidemiology/biostatistics specialty (7.72%). Isfahan, Tehran, Shahid Beheshti, Iran, Baqiyatallah, and Urmia universities of medical sciences had consecutively the largest number of publications in the studied journals. Only three papers (0.39%) were published by the international collaboration. CONCLUSIONS: Iranian nursing journals should publish special issues in the neglected subject areas. These journals should encourage authors to publish research evidence with higher quality.