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The last pregnancy trimester is critical for fetal brain development but is a vulnerable period if the pregnancy is compromised by fetal growth restriction (FGR). The impact of FGR on the maturational development of neuronal morphology is not known, however, studies in fetal sheep allow longitudinal analysis in a long gestation species. Here we compared hippocampal neuron dendritogenesis in FGR and control fetal sheep at three timepoints equivalent to the third trimester of pregnancy, complemented by magnetic resonance image for brain volume, and electrophysiology for synaptic function. We hypothesized that the trajectory of hippocampal neuronal dendrite outgrowth would be decreased in the growth-restricted fetus, with implications for hippocampal volume, connectivity, and function. In control animals, total dendrite length increased with advancing gestation, but not in FGR, resulting in a significantly reduced trajectory of dendrite outgrowth in FGR fetuses for total length, branching, and complexity. Ex vivo electrophysiology analysis shows that paired-pulse facilitation was reduced in FGR compared to controls for cornu ammonis 1 hippocampal outputs, reflecting synaptic dysfunction. Hippocampal brain-derived neurotrophic factor density decreased over late gestation in FGR fetuses but not in controls. This study reveals that FGR is associated with a significant deviation in the trajectory of dendrite outgrowth of hippocampal neurons. Where dendrite length significantly increased over the third trimester of pregnancy in control brains, there was no corresponding increase over time in FGR brains, and the trajectory of dendrite outgrowth in FGR offspring was significantly reduced compared to controls. Reduced hippocampal dendritogenesis in FGR offspring has severe implications for the development of hippocampal connectivity and long-term function., (© 2025 The Author(s). Brain Pathology published by John Wiley & Sons Ltd on behalf of International Society of Neuropathology.)

Fetal growth restriction (FGR) is associated with cardiovascular and respiratory complications after birth and beyond. Despite research showing a range of neurological changes following FGR, little is known about how FGR affects the brainstem cardiorespiratory control centres. The primary neurons that release serotonin reside in the brainstem cardiorespiratory control centres and may be affected by FGR. At two time points in the last trimester of sheep brain development, 110 and 127 days of gestation (0.74 and 0.86 of gestation), we assessed histopathological alterations in the brainstem cardiorespiratory control centres of the pons and medulla in early-onset FGR versus control fetal sheep. The FGR cohort were hypoxaemic and asymmetrically growth restricted. Compared to the controls, the brainstem of FGR fetuses exhibited signs of neuropathology, including elevated cell death and reduced cell proliferation, grey and white matter deficits, and evidence of oxidative stress and neuroinflammation. FGR brainstem pathology was predominantly observed in the medullary raphé nuclei, hypoglossal nucleus, nucleus ambiguous, solitary tract and nucleus of the solitary tract. The FGR groups showed imbalanced brainstem serotonin and serotonin 1A receptor abundance in the medullary raphé nuclei, despite evidence of increased serotonin staining within vascular regions of placentomes collected from FGR fetuses. Our findings demonstrate both early and adaptive brainstem neuropathology in response to placental insufficiency. KEY POINTS: Early-onset fetal growth restriction (FGR) was induced in fetal sheep, resulting in chronic fetal hypoxaemia. Growth-restricted fetuses exhibit persistent neuropathology in brainstem nuclei, characterised by disrupted cell proliferation and reduced neuronal cell number within critical centres responsible for the regulation of cardiovascular and respiratory functions. Elevated brainstem inflammation and oxidative stress suggest potential mechanisms contributing to the observed neuropathological changes. Both placental and brainstem levels of 5-HT were found to be impaired following FGR., (© 2023 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Background: Preclinical evidence indicates that statins possess diverse antineoplastic effects in different types of tumors. However, clinical studies have yielded conflicting results regarding the potential of statins to either increase or decrease the risk of cancer. Our objective was to examine the relationship between the dose of a treatment and its impact on melanoma tumor growth and angiogenesis in an in vivo setting., Materials and Methods: Melanoma cells were injected into C57BL6 mice in four groups. They received 0, 1, 5, and 10 mg/kg of atorvastatin daily. Three others received the mentioned doses one week before the inoculation of melanoma animals. At the end of the third week, the animals were euthanized in a humane manner, and both blood samples and tumor specimens were collected for subsequent analysis., Results: The tumor size was 1.16 ± 0.25 cm 3 in a group treated with therapeutic dose of atorvastatin and was significantly larger than that in the control group (0.42 ± 0.08 cm 3 ). However, there were no significant differences between the two other doses and the control group (0.72 ± 0.22, 0.46 ± 0.08 cm 3 in atorvastatin-treated groups with 5 and 10 mg/kg). The vascular density of the tumors was significantly increased in the lowest dose of the atorvastatin treatment group, similar to the results of tumor size ( P < 0.05)., Conclusion: Atorvastatin, at low therapeutic concentrations, has been observed to stimulate tumor growth and exhibit pro-angiogenic effects. Therefore, it is advised to exercise caution and recommend clinically relevant doses of statins to patients with cancer., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Advanced Biomedical Research.)

Fetal growth restriction (FGR) is a complex obstetric issue describing a fetus that does not reach its genetic growth potential. The primary cause of FGR is placental dysfunction resulting in chronic fetal hypoxaemia, which in turn causes altered neurological, cardiovascular and respiratory development, some of which may be pathophysiological, particularly for neonatal life. The brainstem is the critical site of cardiovascular, respiratory and autonomic control, but there is little information describing how chronic hypoxaemia and the resulting FGR may affect brainstem neurodevelopment. This review provides an overview of the brainstem-specific consequences of acute and chronic hypoxia, and what is known in FGR. In addition, we discuss how brainstem structural alterations may impair functional control of the cardiovascular and respiratory systems. Finally, we highlight the clinical and translational findings of the potential roles of the brainstem in maintaining cardiorespiratory adaptation in the transition from fetal to neonatal life under normal conditions and in response to the pathological environment that arises during development in growth-restricted infants. This review emphasises the crucial role that the brainstem plays in mediating cardiovascular and respiratory responses during fetal and neonatal life. We assess whether chronic fetal hypoxaemia might alter structure and function of the brainstem, but this also serves to highlight knowledge gaps regarding FGR and brainstem development., (© 2023 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Metastasis of melanoma significantly worsens prognosis; thus, therapeutic interventions that prevent metastasis could improve patient outcomes. Here, we show using humanized mice that colonization of distant visceral organs with melanoma is dependent upon a human CD33+CD11b+CD117+ progenitor cell subset comprising <4% of the human CD45+ leukocytes. Metastatic tumor-infiltrating CD33+ cells from patients and humanized (h)NSG-SGM3 mice showed converging transcriptional profiles. Single-cell RNA-seq analysis identified a gene signature of a KIT/CD117-expressing CD33+ subset that correlated with decreased overall survival in a TCGA melanoma cohort. Thus, human CD33+CD11b+CD117+ myeloid cells represent a novel candidate biomarker as well as a therapeutic target for metastatic melanoma., Competing Interests: Disclosures:   C.I. Yu reported a patent to humanized mouse model for cancer metastasis pending. P. Metang reported “other” from University of Texas Southwestern Medical Center outside the submitted work. J. Banchereau reported grants from Merck during the conduct of the study; grants from Sanofi, personal fees from Cue Biopharma, personal fees from Neovacs, personal fees from Ascend Pharma, and personal fees from Georgiamune outside the submitted work; in addition, J. Banchereau had a patent to humanized mouse model for cancer metastasis pending. K. Palucka reported grants from Merck, “other” from Merck, personal fees from Cue Biopharma, and personal fees from Sobi outside the submitted work; in addition, K. Palucka had a patent on humanized mice to study metastasis pending. No other disclosures were reported., (© 2021 Yu et al.)

Thanks to many advances in genetic manipulation, mouse models have become very powerful in their ability to interrogate biological processes. In order to precisely target expression of a gene of interest to particular cell types, intersectional genetic approaches using two promoter/enhancers unique to a cell type are ideal. Within these methodologies, variants that add temporal control of gene expression are the most powerful. We describe the development, validation and application of an intersectional approach that involves three transgenes, requiring the intersection of two promoter/enhancers to target gene expression to precise cell types. Furthermore, the approach uses available lines expressing tTA/rTA to control the timing of gene expression based on whether doxycycline is absent or present, respectively. We also show that the approach can be extended to other animal models, using chicken embryos. We generated three mouse lines targeted at the Tigre ( Igs7 ) locus with TRE- loxP -tdTomato- loxP upstream of three genes ( p21 , DTA and Ctgf ), and combined them with Cre and tTA/rtTA lines that target expression to the cerebellum and limbs. Our tools will facilitate unraveling biological questions in multiple fields and organisms., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2020. Published by The Company of Biologists Ltd.)

Cancer is a leading cause of mortalities worldwide. Over the past few decades, exploration of molecular mechanisms behind cancer initiation and progression has been of great interest in the viewpoint of both basic and clinical scientists. It is generally believed that identification of key molecules implicated in cancer pathology not only improves our understanding of the disease, but also could result in introduction of novel therapeutic strategies. Neutrophil gelatinase-associated lipocalin (NGAL)/lipocalin-2 (LCN2) is a member of lipocalin superfamily with a variety of functions. Although the main function of LCN2 is still unknown, many studies confirmed its significant role in the initiation, progression, and metastasis of various types of cancer. Furthermore, aberrant expression of LCN2 is also concerned with the chemo- and radio-resistant phenotypes of tumors. Here, we will review the contribution of known functions of LCN2 to the pathophysiology of cancer. We also highlight how the deregulated expression of LCN2 is associated with a variety of fatal types of cancer for which there are no effective therapeutic modalities. The unique and multiple functions of LCN2 and its widespread expression in different types of cancer prompted us to suggest LCN2 could be considered either as a valuable diagnostic and prognostic biomarker or as a potential novel therapeutic target.

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) beating can be efficiently characterized by time-lapse quantitative phase imaging (QPIs) obtained by digital holographic microscopy. Particularly, the CM's nucleus section can precisely reflect the associated rhythmic beating pattern of the CM suitable for subsequent beating pattern characterization. In this paper, we describe an automated method to characterize single CMs by nucleus extraction from QPIs and subsequent beating pattern reconstruction and quantification. However, accurate CM's nucleus extraction from the QPIs is a challenging task due to the variations in shape, size, orientation, and lack of special geometry. To this end, we propose a novel fully convolutional neural network (FCN)-based network architecture for accurate CM's nucleus extraction using pixel classification technique and subsequent beating pattern characterization. Our experimental results show that the beating profile of multiple extracted single CMs is less noisy and more informative compared to the whole image slide. Applying this method allows CM characterization at the single-cell level. Consequently, several single CMs are extracted from the whole slide QPIs and multiple parameters regarding their beating profile of each isolated CM are efficiently measured., Competing Interests: The authors declare that there are no conflicts of interest related to this article., (© 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreement.)

Using non-drug, non-surgical treatments for improving bone mineral diseases in newborn babies is an important topic for neonatologists. The present study introduces bacterial synthesized ionic nano-hydroxyapatite (Bio-HA) for the development of bone mineral density in the chicken embryo model. In vitro cytotoxicity analyses were demonstrated the optimal concentrations of Bio-HA compared to a chemically-synthesized hydroxyapatite (Ch-HA). Toxicity of Bio-HA on MCF-7 human cell lines was negligible at the concentrations less than 500 μg/mL whereas Ch-HA showed similar results at the concentrations less than 100 μg/mL. Therefore, concentrations at 50 μg/mL and 100 μg/mL were selected for in ovo injection of both materials into the fertilized eggs. The newly hatched chickens were sacrificed in order to monitor their serological factors, total body mass, bone mineral contents and bone mineral density. The results confirmed that Bio-HA increased the average body weight and bone mineral indices of chickens in comparison to the Ch-HA and negative controls (normal saline and intact groups). In view of the intact group, no liver or kidney damage occurred in the groups receiving Bio-HA which promises the effectiveness of these nanoparticles for the treatment of afterbirth bone mineral deficiency.

This paper investigates the rhythm strip and parameters of synchronization of human induced pluripotent stem cell (iPS) derived cardiomyocytes. The synchronization is evaluated from quantitative phase images of beating cardiomyocytes which are obtained using the time-lapse digital holographic imaging method. By quantitatively monitoring the dry mass redistribution, digital holography provides the physical contraction-relaxation signal caused by autonomous cardiac action potential. In order to analyze the synchronicity at the cell-to-cell level, we extracted single cardiac muscle cells, which contain the nuclei, from the phase images of cardiomyocytes containing multiple cells resulting from the fusion of k-means clustering and watershed segmentation algorithms. We demonstrate that mature cardiomyocyte cell synchronization can be automatically evaluated by time-lapse microscopic holographic imaging. Our proposed method can be applied for studies on cardiomyocyte disorders and drug safety testing systems., Competing Interests: The authors declare that there are no conflicts of interest related to this article.

Cardiomyocytes derived from human pluripotent stem cells are a promising tool for disease modeling, drug compound testing, and cardiac toxicity screening. Bio-image segmentation is a prerequisite step in cardiomyocyte image analysis by digital holography (DH) in microscopic configuration and has provided satisfactory results. In this study, we quantified multiple cardiac cells from segmented 3-dimensional DH images at the single-cell level and measured multiple parameters describing the beating profile of each individual cell. The beating profile is extracted by monitoring dry-mass distribution during the mechanical contraction-relaxation activity caused by cardiac action potential. We present a robust two-step segmentation method for cardiomyocyte low-contrast image segmentation based on region and edge information. The segmented single-cell contains mostly the nucleus of the cell since it is the best part of the cardiac cell, which can be perfectly segmented. Clustering accuracy was assessed by a silhouette index evaluation for k-means clustering and the Dice similarity coefficient (DSC) of the final segmented image. 3D representation of single of cardiomyocytes. The cell contains mostly the nucleus section and a small area of cytoplasm., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We present unsupervised clustering methods for automatic grouping of human red blood cells (RBCs) extracted from RBC quantitative phase images obtained by digital holographic microscopy into three RBC clusters with regular shapes, including biconcave, stomatocyte, and sphero-echinocyte. We select some good features related to the RBC profile and morphology, such as RBC average thickness, sphericity coefficient, and mean corpuscular volume, and clustering methods, including density-based spatial clustering applications with noise, k-medoids, and k-means, are applied to the set of morphological features. The clustering results of RBCs using a set of three-dimensional features are compared against a set of two-dimensional features. Our experimental results indicate that by utilizing the introduced set of features, two groups of biconcave RBCs and old RBCs (suffering from the sphero-echinocyte process) can be perfectly clustered. In addition, by increasing the number of clusters, the three RBC types can be effectively clustered in an automated unsupervised manner with high accuracy. The performance evaluation of the clustering techniques reveals that they can assist hematologists in further diagnosis.

Hydroxyapatite (HA)-coated magnetite nanoparticles (MNPs) are being widely investigated for various applications in medical engineering and wastewater treatment. In this work, the MNPs were thoroughly coated by bacterial synthesized HA nanoparticles during biomineralization process using Enterobacter aerogenes. The resulting bacterial-induced precipitate was then calcined at 600°C and investigated with respect to structural characteristics, particle size and magnetic strength by XRD, FT-IR, SEM, EDS, TEM and VSM analyses. The effects of MNPs and HA-coated MNPs (HA-MNPs) on the viability of human MCF-7 cell lines were also investigated via mitochondrial activity test (MTT) and lactate dehydrogenase (LDH) assays. The powder characterization results showed appropriate structural properties for HA-MNPs samples. The particles diameter size of the MNPs and HA-MNPs were in the range of 3-25nm and 20-80nm, respectively. The biologically-synthesized HA-MNPs formed a stable suspension in water while keeping their magnetic property. The saturation magnetization (Ms) of HA-MNPs was measured at ~10emug -1 which was in good agreement with the structural composition of this sample. Finally, the results of the cell lines viability indicated that coating of toxic MNPs via biomineralization was a promising approach in order to synthesize bio-compatible magnetic nanoparticles with suitable physical and chemical structural characteristics. The toxicity level of MNPs was reduced by 10 fold when coated by bacterial-synthesized HA., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Objectives: Overexpression of ATP-binding cassette (ABC) transporters is a frequent cause of multidrug resistance in cancer cells and pathogenic microorganisms. One example is the Cdr1p transporter from the human fungal pathogen Candida albicans that belongs to the pleiotropic drug resistance (PDR) subfamily of ABC transporters found in fungi and plants. Cdr1p is overexpressed in several azole-resistant clinical isolates, causing azole efflux and treatment failure. Cdr1p appears as a doublet band in western blot analyses, suggesting that the protein is post-translationally modified. We investigated whether Cdr1p is phosphorylated and the function of this modification., Methods: Phosphorylated residues were identified by MS. Their function was investigated by site-directed mutagenesis and expression of the mutants in a C. albicans endogenous system that exploits a hyperactive allele of the Tac1p transcription factor to drive high levels of Cdr1p expression. Fluconazole resistance was measured by microtitre plate and spot assays and transport activity by Nile red accumulation., Results: We identified a cluster of seven phosphorylated amino acids in the N-terminal extension (NTE) of Cdr1p. Mutating all seven sites to alanine dramatically diminished the ability of Cdr1p to confer fluconazole resistance and transport Nile red, without affecting Cdr1p localization. Conversely, a Cdr1p mutant in which the seven amino acids were replaced by glutamate was able to confer high levels of fluconazole resistance and to export Nile red., Conclusions: Our results demonstrate that the NTE of Cdr1p is phosphorylated and that NTE phosphorylation plays a major role in regulating Cdr1p and possibly other PDR transporter function., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

To repair damaged bone tissues, osteoconductive bone graft substitutes are required for enhancement of the regenerative potential of osteoblast cells. Nanostructured hydroxyapatite is a bioactive ceramic used for bone tissue engineering purposes. In this study, carbonate hydroxyapatite (cHA) and zinc-magnesium substituted hydroxyapatite (Zn-Mg-HA) nanoparticles were synthesized via biomineralization method using Enterobacter aerogenes. The structural phase composition and the morphology of the samples were analyzed using appropriate powder characterization methods. Next, a composite graft was fabricated by using polyvinyl alcohol and both cHA and Zn-Mg-HA samples. In vivo osteogenic potential of the graft was then investigated in a rabbit tibial osteotomy model. Histological, radiological and morphological studies showed that the graft was mineralized by the newly formed bone tissue without signs of inflammation or infection after 4 weeks of implantation. These histomorphometric results suggest that the fabricated graft can function as a potent osteoconductive bone tissue substitute., (Copyright © 2016 Elsevier Inc. All rights reserved.)