Search Results (13)

Solid organ transplantation remains a life-saving treatment for patients worldwide. Unfortunately, the supply of donor organs cannot meet the current need, making the search for alternative sources even more essential. Xenotransplantation using sophisticated genetic engineering techniques to delete and overexpress specific genes in the donor animal has been investigated as a possible option. However, the use of exogenous tissue presents another host of obstacles, particularly regarding organ rejection. Given these limitations, interspecies blastocyst complementation in combination with precise gene knockouts presents a unique, promising pathway for the transplant organ shortage. In recent years, great advancements have been made in the field, with encouraging results in producing a donor-derived organ in a chimeric host. That said, one of the major barriers to successful interspecies chimerism is the mismatch in the developmental stages of the donor and the host cells in the chimeric embryo. Another major barrier to successful chimerism is the mismatch in the developmental speeds between the donor and host cells in the chimeric embryos. This review outlines 19 studies in which blastocyst complementation was used to generate solid organs. In particular, the genesis of the liver, lung, kidney, pancreas, heart, thyroid, thymus and parathyroids was investigated. Of the 19 studies, 7 included an interspecies model. Of the 7, one was completed using human donor cells in a pig host, and all others were rat–mouse chimeras. While very promising results have been demonstrated, with great advancements in the field, several challenges continue to persist. In particular, successful chimerism, organ generation and donor contribution, synchronized donor–host development, as well as ethical concerns regarding human–animal chimeras remain important aspects that will need to be addressed in future research. Full article

Multitarget compounds have emerged as promising drug candidates to cope with complex multifactorial diseases, like Alzheimer’s disease (AD). Most multitarget compounds are designed by linking two pharmacophores through a tether chain (linked hybrids), which results in rather large molecules that are particularly useful to hit targets with large binding cavities, but at the expense of suffering from suboptimal physicochemical/pharmacokinetic properties. Molecular size reduction by removal of superfluous structural elements while retaining the key pharmacophoric motifs may represent a compromise solution to achieve both multitargeting and favorable physicochemical/PK properties. Here, we report the stepwise structural simplification of the dihydroxyanthraquinone moiety of a rhein–huprine hybrid lead by hydroxy group removal—ring contraction—ring opening—ring removal, which has led to new analogs that retain or surpass the potency of the lead on its multiple AD targets while exhibiting more favorable drug metabolism and pharmacokinetic (DMPK) properties and safety profile. In particular, the most simplified acetophenone analog displays dual nanomolar inhibition of human acetylcholinesterase and butyrylcholinesterase (IC₅₀ = 6 nM and 13 nM, respectively), moderately potent inhibition of human BACE-1 (48% inhibition at 15 µM) and Aβ42 and tau aggregation (73% and 68% inhibition, respectively, at 10 µM), favorable in vitro brain permeation, higher aqueous solubility (18 µM) and plasma stability (100/96/86% remaining in human/mouse/rat plasma after 6 h incubation), and lower acute toxicity in a model organism (zebrafish embryos; LC₅₀ >> 100 µM) than the initial lead, thereby confirming the successful lead optimization by structural simplification. Full article

Despite the extensive information available on the different genetic, epigenetic, and molecular features of cardiogenesis, the origin of congenital heart defects remains unknown. Most genetic and molecular studies have been conducted outside the context of the progressive anatomical and histological changes in the embryonic heart, which is one of the reasons for the limited knowledge of the origins of congenital heart diseases. We integrated the findings of descriptive studies on human embryos and experimental studies on chick, rat, and mouse embryos. This research is based on the new dynamic concept of heart development and the existence of two heart fields. The first field corresponds to the straight heart tube, into which splanchnic mesodermal cells from the second heart field are gradually recruited. The overall aim was to create a new vision for the analysis, diagnosis, and regionalized classification of congenital defects of the heart and great arteries. In addition to highlighting the importance of genetic factors in the development of congenital heart disease, this study provides new insights into the composition of the straight heart tube, the processes of twisting and folding, and the fate of the conus in the development of the right ventricle and its outflow tract. The new vision, based on in vivo labeling and cell tracking and enhanced by models such as gastruloids and organoids, has contributed to a better understanding of important errors in cardiac morphogenesis, which may lead to several congenital heart diseases. Full article

Improved genome editing via oviductal nucleic acids delivery (i-GONAD) is a new technology enabling in situ genome editing of mammalian zygotes exiting the oviductal lumen, which is now available in mice, rats, and hamsters. In this method, CRISPR/Cas9 genome-editing reagents are delivered directly to the oviducts of pregnant animals (corresponding to late zygote stage). After intraoviductal instillation, electric shock to the entire oviduct was provided with a specialized electroporation (EP) device to drive the genome editing reagents into the zygotes present in the oviductal lumen. i-GONAD toward early zygotes has been recognized as difficult, because they are tightly surrounded by a cumulus cell layer, which often hampers effective transfer of nucleic acids to zygotes. However, in vivo EP three min after intraoviductal instillation of the genome-editing reagents enabled genome editing of early zygotes with an efficiency of 70%, which was in contrast with the rate of 18% when in vivo EP was performed immediately after intraoviductal instillation at Day 0.5 of pregnancy (corresponding to 13:00–13:30 p.m. on the day when vaginal plug was recognized after natural mating). We also found that addition of hyaluronidase, an enzyme capable of removing cumulus cells from a zygote, slightly enhanced the efficiency of genome editing in early zygotes. These findings suggest that cumulus cells surrounding a zygote can be a barrier for efficient generation of genome-edited mouse embryos and indicate that a three-minute interval before in vivo EP is effective for achieving i-GONAD-mediated genome editing at the early zygote stage. These results are particularly beneficial for researchers who want to perform genome editing experiments targeting early zygotes. Full article

CYP1A is a heme-thiolate enzyme associated with the cytochrome P4501A1 monooxygenase system and is inducible by a wide variety of xenobiotics and endogenous ligands that bind and activate the aryl hydrocarbon receptor (AHR). The AHR-CYP1A axis is important for detoxification of certain xenobiotics and for homeostatic balance of endogenous sex hormones, amine hormones, vitamins, fatty acids, and phospholipids. Herein, we generated and described applications of a zebrafish CYP1A-targeted monoclonal antibody (mAb CRC4) that fortuitously recognizes induced CYP1A across vertebrate taxa, including fish, chicken, mouse, rat, and human. We then demonstrated that mAb CRC4 targets a highly conserved epitope signature of vertebrate CYP1A. The unique complimentary determining region (CDR) sequences of heavy and light chains were determined, and these Ig sequences will allow for the expression of recombinant mAb CRC4, thus superseding the need for long-term hybridoma maintenance. This antibody works well for immunohistochemistry (IHC), as well as whole-mounted IHC in zebrafish embryos. Monoclonal antibody CRC4 may be particularly useful for studying the AHR-CYP1A axis in multiple vertebrate species and within the context of Oceans and Human Health research. By using archived samples, when possible, we actively promoted efforts to reduce, replace, and refine studies involving live animals. Full article

The purpose of this study was to evaluate the potential of a newly modified cyclodextrin derivative, water-soluble β-cyclodextrin–epichlorohydrin (β-CD), as an effective drug carrier to enhance the poor solubility and bioavailability of galangin (GAL), a poorly water-soluble model drug. In this regard, inclusion complexes of GAL/β-CDP were prepared. UV-VIS spectrophotometry, Fourier-transform infrared spectroscopy (FTIR), X-ray crystallography (XRD), zeta potential analysis, particle size analysis, field emission scanning electron microscopy (FESEM), and transmission electron microscopy (TEM) were applied to characterize the synthesized GAL/β-CD. Michigan Cancer Foundation-7 (MCF-7; human breast cancer cells) and rat embryo fibroblast (REF; normal cells) were employed to examine the in vitro cytotoxic effects of GAL/β-CD using various parameters. The dye-based tests of MTT and crystal violet clearly exhibited that GAL/β-CD-treated cells had a reduced proliferation rate, an influence that was not found in the normal cell line. The cells’ death was found to follow apoptotic mechanisms, as revealed by the dye-based test of acridine orange/ethidium bromide (AO/EtBr), with the involvement of the mitochondria via caspase-3-mediated events, as manifested by the Rh 123 test. We also included a mouse model to examine possible in vivo toxic effects of GAL/β-CD. It appears that the inclusion complex does not have a significant influence on normal cells, as indicated by serum levels of kidney and liver enzymatic markers, as well as thymic and splenic mass indices. A similar conclusion was reached on the histological level, as manifested by the absence of pathological alterations in the liver, kidney, thymus, spleen, heart, and lung. Full article

Kidney disease patients may have concurrent chronic kidney disease-associated mineral bone disorder and hypertension. Cardiovascular disease (CVD) and neuropathy occur due to kidney failure-induced accumulation of uremic toxins in the body. Indoxyl sulfate (IS), a product of indole metabolism in the liver, is produced from tryptophan by the intestinal flora and is ultimately excreted through the kidneys. Hemodialysis helps renal failure patients eliminate many nephrotoxins, except for IS, which leads to a poor prognosis. Although the impacts of IS on cardiac and renal development have been well documented using mouse and rat models, other model organisms, such as zebrafish, have rarely been studied. The zebrafish genome shares at least 70% similarity with the human genome; therefore, zebrafish are ideal model organisms for studying vertebrate development, including renal development. In this study, we aimed to investigate the impact of IS on the development of zebrafish embryos, especially cardiac and renal development. At 24 h postfertilization (hpf), zebrafish were exposed to IS at concentrations ranging from 2.5 to 10 mM. IS reduced survival and the hatching rate, caused cardiac edema, increased mortality, and shortened the body length of zebrafish embryos. In addition, IS decreased heart rates and renal function. IS affected zebrafish development via the ROS and MAPK pathways, which subsequently led to inflammation in the embryos. The results suggest that IS interferes with cardiac and renal development in zebrafish embryos, providing new evidence about the toxicity of IS to aquatic organisms and new insights for the assessment of human health risks. Accordingly, we suggest that zebrafish studies can ideally complement mouse model studies to allow the simultaneous and comprehensive investigation of the physiological impacts of uremic endotheliotoxins, such as IS, on cardiac and renal development. Full article

Omega-3 polyunsaturated fatty acids (n-3 PUFAs) play an important role in the development, maintenance, and function of the brain. Dietary supplementation of n-3 PUFAs in neurological diseases has been a subject of particular interest in preventing cognitive deficits, and particularly in age-related neurodegeneration. Developing strategies for the efficient delivery of these lipids to the brain has presented a challenge in recent years. We recently reported the preparation of n-3 PUFA-rich nanoliposomes (NLs) from salmon lecithin, and demonstrated their neurotrophic effects in rat embryo cortical neurons. The objective of this study was to assess the ability of these NLs to deliver PUFAs in cellulo and in vivo (in mice). NLs were prepared using salmon lecithin rich in n-3 PUFAs (29.13%), and characterized with an average size of 107.90 ± 0.35 nm, a polydispersity index of 0.25 ± 0.01, and a negative particle-surface electrical charge (−50.4 ± 0.2 mV). Incubation of rat embryo cortical neurons with NLs led to a significant increase in docosahexaenoic acid (DHA) (51.5%, p < 0.01), as well as palmitic acid, and a small decrease in oleic acid after 72 h (12.2%, p < 0.05). Twenty mice on a standard diet received oral administration of NLs (12 mg/mouse/day; 5 days per week) for 8 weeks. Fatty acid profiles obtained via gas chromatography revealed significant increases in cortical levels of saturated, monounsaturated, and n-3 (docosahexaenoic acid,) and n-6 (docosapentaenoic acid and arachidonic acid) PUFAs. This was not the case for the hippocampus or in the liver. There were no effects on plasma lipid levels, and daily monitoring confirmed NL biocompatibility. These results demonstrate that NLs can be used for delivery of PUFAs to the brain. This study opens new research possibilities in the development of preventive as well as therapeutic strategies for age-related neurodegeneration. Full article

Background: Hypertensive disorders of pregnancy, gestational hypertension, and preeclampsia (PE) are exceptionally challenging, as their pathologies and therapeutic management simultaneously influence the mother and embryo, sometimes putting their well-beings at odds with each other. Dysregulated lipid and glucose metabolism may be related to some cases of preeclampsia. Fluctuations in serum apelin levels may be attributed to changes in the serum levels of multiple interrelated factors such as insulin, insulin resistance, inflammatory cytokines, and nephritic damage. Previous studies demonstrated that apelin is an endogenous active peptide with vasodilatory and antioxidative-stress capabilities. Objective: We investigated the relationships among hepatic, nephrotic, and metabolic injuries in different preeclampsia-like mouse models and the potential effect of exogenous apelin administration on hepatic and nephrotic injuries and metabolic disorders in an N-nitro-L-arginine methyl ester (L-NAME) preeclampsia-like Sprague Dawley (SD) rat model. Materials and methods: Forty-three adult female and ten adult male SD rats were involved in this study. The male rats were used to induce pregnancy. The female rats were randomly divided into four equal groups: a non-pregnant group, a normal pregnant group, a group treated with L-NAME to induce preeclampsia, and a group treated with L-NAME and apelin. In all the groups, maternal blood was collected on the 21st day of gestation, and serum samples were used for the determination of systolic blood pressure; the serum uric acid, creatinine, nitric oxide (NO), xanthine oxidase, myeloperoxidase, insulin, glucose, cholesterol, triglyceride (TG), aspartate aminotransferase (AST) and alanine aspartate aminotransferase (ALT) levels; and the HOMA-insulin resistance (HOMA-IR). Results: In rats with pre-eclampsia, the systolic blood pressure; the concentrations of serum uric acid, creatinine, nitric oxide (NO), xanthine oxidase, myeloperoxidase, blood glucose, serum cholesterol, triglycerides, AST, and ALT; and the calculated HOMA-IR were significantly increased compared to those in the rats in the other groups. Additionally, apelin treatment significantly improved these parameters in the apelin-treated group. Conclusion: This study examined the potential mechanisms whereby apelin may act as a curative candidate to reduce hepatic injury and inhibit kidney damage and the development of metabolic disorders in an experimental model of preeclampsia. Full article

Since its discovery in Drosophila, the Notch signaling pathway has been studied in numerous developmental contexts in diverse multicellular organisms. The role of Notch signaling in nervous system development has been extensively investigated by numerous scientists, partially because many of the core Notch signaling components were initially identified through their dramatic ‘neurogenic’ phenotype of developing fruit fly embryos. Components of the Notch signaling pathway continue to be expressed in mature neurons and glia cells, which is suggestive of a role in the post-developmental nervous system. The Notch pathway has been, so far, implicated in learning and memory, social behavior, addiction, and other complex behaviors using genetic model organisms including Drosophila and mice. Additionally, Notch signaling has been shown to play a modulatory role in several neurodegenerative disease model animals and in mediating neural toxicity of several environmental factors. In this paper, we summarize the knowledge pertaining to the post-developmental roles of Notch signaling in the nervous system with a focus on discoveries made using the fruit fly as a model system as well as relevant studies in C elegans, mouse, rat, and cellular models. Since components of this pathway have been implicated in the pathogenesis of numerous psychiatric and neurodegenerative disorders in human, understanding the role of Notch signaling in the mature brain using model organisms will likely provide novel insights into the mechanisms underlying these diseases. Full article

During the last five decades, a large number of BT (Benzothiazole) derivatives formed one of the eligible structures in medicinal chemistry as anticancer agents. Most of the studies reveal that various substitutions at specific positions on BT scaffold modulate the antitumor property. The potential of BTs encouraged us to synthesize a number of new 2-((5-substitutedbenzothiazol-2-yl)thio)-N’-(2-(4-(substitutedphenyl)ethylidene)acetohydrazide derivatives and investigate their probable anticancer activity. 4-Substitued benzaldehyde derivatives (1a–1e) were afforded by the reaction of appropriate secondary amine and 4-fluorobenzaldehyde in DMF. Equimolar quantitates of 5-substitutedbenzothiazole-2-thiol, ethyl chloroacetate and K₂CO₃ were refluxed in acetone to obtain 2-((5-substitutedbenzothiazol-2-yl)thio)acetate derivatives (2a,2b), which reacted with excess of hydrazine hydrate to get 2-((5-substitutebenzothiazol-2-yl)thio)acetohydrazides (3a,3b). In the last step, 2-((5-substitutedbenzothiazol-2-yl)thio)-N’-(4-substitutedbenzylidene)acetohydrazide derivatives (4a–4j) were synthesized by the reaction of 1a–1e and 3a–3b in EtOH. The anticancer activity of target compounds was evaluated in three steps. First, an MTT test (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) was performed to observe cytotoxic activity of the compounds against carcinogenic C6 (Rat brain glioma cell line), A549 (Human lung adenocarcinoma epithelial cell line), MCF-7 (Human breast adenocarcinoma cell line), and HT-29 (Human colorectal adenocarcinoma cell line) cancer cell lines. Healthy NIH3T3 (Mouse embryo fibroblast cell line) cells were also subjected to MTT assay to determine selectivity of the compounds towards carcinogenic cell lines. Secondly, inhibitory effects of selected compounds 4d, 4e, and 4h on DNA synthesis of C6 cells were investigated. Finally, flow cytometric analysis were performed to identify the death pathway of the carcinogenic cells. Full article

Currently in vitro culture of mouse preimplantation embryos has become a very important technique to investigate different mechanisms of early embryogenesis. However, there is a big difference in the preimplantation development between mammalian species. Despite close relatedness to mice, in vitro cultivation of rat preimplantation embryos is still delicate and needs further investigation and optimizations. In this study we have compared the in vitro developmental potential of mouse and rat embryos cultured at different culture conditions in parallel experiments. Interestingly, mouse zygotes developed in vitro until blastocyst stage even in inadequate medium without any phosphates and with low osmolarity which was formulated especially for cultivation of rat embryos. Rat parthenotes and zygotes developed in M16 medium formulated for mouse embryos only till 2-cell stage and further development is blocked completely at this stage. Moreover, developmental ability of rat embryos in vitro was significantly lower in comparison with mouse even in special rat mR1ECM medium. Mouse and rat embryos at 2-cell stage obtained in vivo developed until blastocyst stages significantly more efficiently compared to zygotes. Culture of mouse zygotes in glass capillaries resulted in a significantly higher rate of morula and blastocyst development compared with dishes. The Well-of-the-Well system resulted in a significant improvement when compared with dishes for the culture of rat zygotes only until morula stage. Reduced oxygen tension increased the developmental rate of rat but not mouse zygotes until blastocyst stage. This study demonstrates that development of early preimplantation embryos is altered by different culture conditions and show strong differences even between two related species such as mice and rats. Therefore, for understanding the fundamental mechanisms of early mammalian development it is very important to use embryos of various species. Full article

The implantation process is complex, requiring reciprocal interactions between implantation-competent blastocysts and the receptive uterus. There were reports to show that some microRNAs (miRNAs) may play a key role during embryo implantation in mouse. However, the miR-320 expression profiles in the rat uterus during peri-implantation are unknown. In the present study, we found that the expression level of miR-320 was lower on day 5 of gestation (g.d. 5) in rats than g.d.3 and g.d.4 and restored gradually from g.d.6. MiR-320 was specifically localized in glandular and luminal epithelia and decidua. The expression of miR-320 was not significantly different in the pseudopregnant uterus and decreased in the uteri of rats subjected to activation of delayed implantation. Artificial decidualization and treatment with progesterone increased the miR-320 expression. Thus, miR-320 was differentially expressed in the rat uterus during implantation. The expression level was affected by active blastocysts and decidualization during the window of implantation. Steroid hormones, progesterone stimulated miR-320 expression. Full article