CD-1 Mouse Model Research Articles

O-038 Impact of maternal aging on the DNA repair landscape in mouse oocytes

Abstract Study question How does maternal aging impact DNA damage accumulation and DNA repair capacity in mammalian oocytes? Summary answer Age-related changes in DNA repair compartments and chromatin organisation drive DNA damage accumulation in aged oocytes, ultimately contributing to the decline in DNA repair capacity. What is known already Oocyte loss occurs progressively from birth and accelerates as women approach their mid-30s, posing a significant challenge to female reproductive health. Recent studies have highlighted the role of the DNA repair response in driving ovarian ageing and oocyte loss. Aged oocytes accumulate elevated levels of DNA damage. Unless repaired, persistent DNA damage activates DNA repair checkpoints, causing the elimination of defective oocytes. Although we know DNA double-strand breaks (DSBs) in oocytes increase with maternal age, surprisingly, little is known about why this damage is not repaired efficiently. Study design, size, duration Oocytes were isolated from ovaries of 8 weeks or 68-86-week-old CD1 mice, which correspond to reproductively young and old females respectively. Over 600 and 1000 oocytes from aged any young females were used to analyse DNA damage levels, DNA repair effciency, DNA repair compartments and for other live assays. Participants/materials, setting, methods DNA damage was induced in CD1 oocytes from young and aged mice using neocarzinostatin. Levels of DNA damage and DNA repair proteins were quantified using immunofluoresnce confocal/STED microscopy. High-resolution light sheet microscopy was used to perform live imaging of dynamics of DNA damage foci and chromosome segregation during meiosis. Main results and the role of chance Our work demonstrates that aged oocytes have a lower DNA repair capacity and reduced mobility of DNA damage sites compared to young oocytes. We further show that incomplete DNA repair results in defective chromosome integrity and partitioning, thereby compromising oocyte quality. To investigate the causes of the age-related decline in DNA repair, we systematically studied the localization and abundance of several DNA repair proteins in oocytes from young and aged mice. We found that DNA repair proteins localized in spatially distinct DNA repair compartments in oocyte nuclei, that form during the late stages of oocyte growth, accompanied by changes in the activity of DNA repair pathways. Several DNA repair proteins, their corresponding compartments, and their response to DNA damage were altered in aged oocytes. Lastly, we demonstrate that age-related cohesin loss increases DNA damage levels and reduces DNA repair capacity. We hypothesize that cohesin dissociation with maternal ageing may make chromosomes prone to accumulate DNA damage without proper DNA repair. Thus, age-related changes in DNA repair compartments and chromatin organisation drive DNA damage accumulation in aged oocytes, ultimately contributing to the decline in reproductive potential. Limitations, reasons for caution Neocarzinostatin, a radiomimetic drug used to induce DNA DSBs in mouse oocytes in vitro, may not fully represent sources of DNA damage during aging. The CD1 mouse model and ages were selected to recapitulate equivalent advanced maternal age in women. Further investigation is needed to confirm these findings in humans. Wider implications of the findings Overall, we propose that altered DNA repair machinery and cohesin loss contribute to persistent DNA damage and reduced DNA repair capacity in aged oocytes, which can cause chromosome defects and fragmentation, and eventually trigger oocyte death, thereby accelerating the loss of female fertility. Trial registration number Not Applicable

Maternal exposure to beta-Cypermethrin disrupts placental development by dysfunction of trophoblast cells from oxidative stress

As a broad-spectrum and efficient insecticide, beta-Cypermethrin (β-CYP) poses a health risk to pregnancy. It matters the mechanisms of maternal exposure to β-CYP for impacting reproductive health. The placenta, a transient organ pivotal for maternal-fetal communication during pregnancy, plays a crucial role in embryonic development. The effect of β-CYP exposure on the placenta and its underlying molecular mechanisms remain obscure. The objective of this study was to investigate the effect of β-CYP exposure on placental development and the function of trophoblast, as well as the underlying mechanisms through CD-1 mouse model (1, 10, 20 mg/kg.bw) and in vitro HTR-8/SVneo cell model (12.5, 25, 50, 100 μM). We found slower weight gain and reduced uterine wet weight in pregnant mice with maternal exposure to β-CYP during pregnancy, as well as adverse pregnancy outcomes such as uterine bleeding and embryo resorption. The abnormal placental development in response to β-CYP was noticed, including imbalanced placental structure and disrupted labyrinthine vascular development. Trophoblasts, pivotal in placental development and vascular remodeling, displayed abnormal differentiation under β-CYP exposure. This aberration was characterized by thickened trophoblast layers in the labyrinthine zone, accompanied by mitochondrial and endoplasmic reticulum swelling within trophoblasts. Further researches on human chorionic trophoblast cell lines revealed that β-CYP exposure induced apoptosis in HTR-8/SVneo cells. This induction resulted in a notable decrease in migration and invasion abilities, coupled with oxidative stress and the inhibition of the Notch signaling pathway. N-acetylcysteine (an antioxidant) partially restored the impaired Notch signaling pathway in HTR-8/SVneo cells, and mitigated cellular functional damage attributed to β-CYP exposure. Collectively, exposure to β-CYP induced oxidative stress and then led to inhibition of the Notch signaling pathway and dysfunction of trophoblast cells, ultimately resulted in abnormal placenta and pregnancy. These findings indicate Reactive Oxygen Species as potential intervention targets to mitigate β-CYP toxicity. The comprehensive elucidation contributes to our understanding of β-CYP biosafety and offers an experimental basis for preventing and managing its reproductive toxicity.

Investigating the effects of valproic acid on placental epigenetic modifications and development in the CD-1 mouse model

Gestational exposure to the anticonvulsant drug valproic acid (VPA) is associated with congenital malformations and neurodevelopmental disorders through its action as a histone deacetylase inhibitor. VPA can elicit placental toxicity and affect placental growth and development. The objective of this study was to evaluate the impact of maternal exposure to VPA on the mouse placenta following exposure on gestational day (GD) 13 since previous studies have shown that mice exposed at this time during gestation give birth to offspring with an autism spectrum disorder-like phenotype. We exposed CD-1 dams to a teratogenic dose (600 mg/kg) of VPA or saline on GD13 and assessed fetoplacental growth and development on GD18. We evaluated epigenetic modifications, including acetylated histone H4 (H4ac), methylated H3K4 (H3K4me2) using immunohistochemistry, and global DNA methylation in the placenta at 1, 3, and 24 h following maternal exposure on GD13. In utero exposure to VPA on GD13 significantly decreased placental weight and increased fetal resorptions. Moreover, VPA significantly increased the staining intensity of histone H4 acetylation and H3K4 di-methylation across the placenta at 1 and 3 h post maternal dose. Our results also demonstrate that VPA significantly decreased global DNA methylation levels in placental tissue. These results show that gestational exposure to VPA interferes with placental growth and elicits epigenetic modifications, which may play a vital role in VPA-induced developmental toxicity.

Intranasal challenge with B. pertussis leads to more severe disease manifestations in mice than aerosol challenge.

The murine Bordetella pertussis challenge model has been utilized in preclinical research for decades. Currently, inconsistent methodologies are employed by researchers across the globe, making it difficult to compare findings. The objective of this work was to utilize the CD-1 mouse model with two routes of challenge, intranasal and aerosol administration of B. pertussis, to understand the differences in disease manifestation elicited via each route. We observed that both routes of B. pertussis challenge result in dose-dependent colonization of the respiratory tract, but overall, intranasal challenge led to higher bacterial burden in the nasal lavage, trachea, and lung. Furthermore, high dose intranasal challenge results in induction of leukocytosis and pro-inflammatory cytokine responses compared to aerosol challenge. These data highlight crucial differences in B. pertussis challenge routes that should be considered during experimental design.

Main Cardiac Histopathologic Alterations in the Acute Phase of Trypanosoma cruzi Infection in a Murine Model.

Symptoms in the acute phase of Chagas disease are usually mild and nonspecific. However, after several years, severe complications like dilated heart failure and even death may arise in the chronic phase. Due to the lack of specific symptoms in the acute phase, the aim of this work was to describe and analyze the cardiac histopathology during this phase in a CD1 mouse model by assessing parasitism, fibrotic damage, and the presence and composition of a cellular infiltrate, to determine its involvement in the pathogenesis of lesions in the cardiac tissue. Our results indicate that the acute phase lasts about 62 days post-infection (dpi). A significant increase in parasitemia was observed since 15 dpi, reaching a maximum at 33 dpi (4.1 × 106). The presence of amastigote nests was observed at 15-62 dpi, with a maximum count of 27 nests at 35 dpi. An infiltrate consisting primarily of macrophages and neutrophils was found in the cardiac tissue within the first 30 days, but the abundance of lymphocytes showed an 8 ≥ fold increase at 40-62 dpi. Unifocal interstitial fibrosis was identified after 9 dpi, which subsequently showed a 16 ≥ fold increase at 40-60 dpi, along with a 50% mortality rate in the model under study. The increased area of fibrotic lesions revealed progression in the extent of fibrosis, mainly at 50-62 dpi. The presence of perivasculitis and thrombus circulation disorders was seen in the last days (62 dpi); finally, cases of myocytolysis were observed at 50 and 62 dpi. These histopathological alterations, combined with collagen deposition, seem to lead to the development of interstitial fibrosis and damage to the cardiac tissue during the acute phase of infection. This study provides a more complete understanding of the patterns of histopathological abnormalities involved in the acute phase, which could help the development of new therapies to aid the preclinical tests of drugs for their application in Chagas disease.

BDNF gene delivery to the retina by cell adhesion peptide-conjugated Gemini Nanoplexes in vivo.

Retinal ganglion cell (RGC) neurodegeneration in glaucoma is not prevented by controlling the elevated intraocular pressure alone. Neuroprotective gene therapy approaches could be an essential part of a combination treatment. Five cell adhesion peptide (CAP)-gemini surfactants (18-7 N(p1-5)-18) were synthesized as building blocks for BDNF gene carrier nanoparticles (CAP-NPXs). The composition of CAP-NPXs was optimized, physicochemically characterized and evaluated for in vitro transfection efficiency (TE) in A7 astrocytes and 3D retinal neurospheres; and for gene expression in vivo in CD1 mice using RFP reporter gene and BDNF levels after intravitreal (IVT) injection. The IgSF-binding 18-7 N(pFASNKL)-18 pNPXs treated cells demonstrated 1.4-fold higher TE compared to integrin-binding 18-7 N(pRGD)-18 pNPXs and parent 18-7NH-18 NPXs with overall viability between 86 and 95%. The 18-7 N(pFASNKL)-18 pNPXs selectively transfected RGCs in 3D MiEye8 neurospheres. In the in vivo CD1 mouse model 18-7 N(pFASNKL)-18 pNPXs administered by IVT injection delivered tdTomato/BDNF plasmid to retinal cells and produced higher gene expression than the 18-7 N(pRGD)-18 pNPXs, the parent 18-7NH-18 NPXs and Lipofectamine® 3000 as demonstrated by confocal microscopy of whole mount retinas. The BDNF gene expression, assessed by ELISA, showed significantly high levels of BDNF with 18-7 N(pFASNKL)-18 (422.60 ± 42.60 pg/eye), followed by 18-7 N(pRGD)-18 pNPXs (230.62 ± 24.47 pg/eye), 18-7NH-18 NPXs (245.90 ± 39.72 pg/eye), Lipofectamine® 3000 (199.99 ± 29.90 pg/eye) and untreated controls (131.33 ± 20.30 pg/eye). In sum, the 18-7 N(pFASNKL)-18 pNPXs induced 3.4-fold higher BDNF level compared to controls and 2-fold higher than 18-7 N(pRGD)-18 pNPXs. The in vivo efficacy of 18-7 N(pFASNKL)-18 NPXs to produce BDNF levels at pharmacologically relevant levels supports further studies.

Fermented Sprouts of Codonopsis lanceolata Suppress LPS-Induced Inflammatory Responses by Inhibiting NF-κB Signaling Pathway in RAW 264.7 Macrophages and CD1 Mice.

The interest in bioconversion through fermentation of sprouts produced in smart farms is increasing due to their potential health benefits. Codonopsis lanceolata (CL) is reported to alleviate inflammatory conditions, but much research is still needed to determine which types and parts of CL are most effective. This study investigated the anti-inflammatory effects of a fermented extract of CL sprouts' aerial part (F-CSA) against LPS-stimulated RAW 264.7 macrophages and mice. In the screening test, F-CSA showed the most substantial anti-inflammatory effect among several samples, containing the highest total flavonoids, tannins, and polyphenols. UPLC-ESI-Q/TOF-MS and HPLC analysis revealed that F-CSA had the highest amount of luteolin among all the CL samples analyzed. F-CSA reduced the release of inflammatory cytokines and mediators such as NO and PGE2 by inhibiting the expression levels of iNOS and COX-2 in LPS-stimulated macrophages. Further, we found that the anti-inflammatory effects of F-CSA were mediated by inhibiting the JNK/NF-κB signaling pathway. Moreover, F-CSA improved survival rates and reduced plasma levels of NO and IL-6 in CD1 mice stimulated with LPS. These findings suggest that F-CSA, which contains luteolin, can alleviate inflammation in LPS-induced RAW 264.7 cells and a CD1 mouse model by inhibiting the JNK/NF-κB signaling pathways.

Single-Cell RNA-Sequencing Reveals Interactions between Endometrial Stromal Cells, Epithelial Cells, and Lymphocytes during Mouse Embryo Implantation

The decidualization of endometrial stromal cells (ESCs) is an essential process facilitating embryo implantation. However, the roles of non-decidualized and decidualized ESCs in regulating the microenvironment of a receptive endometrium remain unclear. We investigated single-cell transcriptomic changes in the uterus of a CD-1 mouse model at the post-implantation stage. The implantation and inter-implantation sites of the uteruses of pregnant mice at 4.5 and 5.5 days post-coitum were dissected for single-cell RNA sequencing. We identified eight cell types: epithelial cells, stromal cells, endothelial cells, mesothelial cells, lymphocytes, myocytes, myeloids, and pericytes. The ESC transcriptome suggests that the four ESC subtypes are involved in the extracellular remodeling during implantation. The trajectory plot of ESC subtypes indicates embryo implantation that involves a differentiation pathway from undifferentiated ESCs (ESC 1) to decidualized ESCs (DEC ESCs), with distinct signaling pathways between the ESC subtypes. Furthermore, the ligand-receptor analysis suggests that ESCs communicate with epithelial cells and immune cells through nectin and ICAM signaling. Collectively, both decidualized and non-decidualized ESCs may regulate the endometrial microenvironment for optimal endometrial receptivity and immune tolerance. This study provides insights on the molecular and cellular characteristics of mouse ESCs in modulating the epithelial and lymphocyte functions during early embryo implantation.

Long-Term Analysis of Pertussis Vaccine Immunity to Identify Potential Markers of Vaccine-Induced Memory Associated With Whole Cell But Not Acellular Pertussis Immunization in Mice.

Over two decades ago acellular pertussis vaccines (aP) replaced whole cell pertussis vaccines (wP) in several countries. Since then, a resurgence in pertussis has been observed, which is hypothesized to be linked, in part, to waning immunity. To better understand why waning immunity occurs, we developed a long-term outbred CD1 mouse model to conduct the longest murine pertussis vaccine studies to date, spanning out to 532 days post primary immunization. Vaccine-induced memory results from follicular responses and germinal center formation; therefore, cell populations and cytokines involved with memory were measured alongside protection from challenge. Both aP and wP immunization elicit protection from intranasal challenge by decreasing bacterial burden in both the upper and lower airways, and by generation of pertussis specific antibody responses in mice. Responses to wP vaccination were characterized by a significant increase in T follicular helper cells in the draining lymph nodes and CXCL13 levels in sera compared to aP mice. In addition, a population of B. pertussis + memory B cells was found to be unique to wP vaccinated mice. This population peaked post-boost, and was measurable out to day 365 post-vaccination. Anti-B. pertussis and anti-pertussis toxoid antibody secreting cells increased one day after boost and remained high at day 532. The data suggest that follicular responses, and in particular CXCL13 levels in sera, could be monitored in pre-clinical and clinical studies for the development of the next-generation pertussis vaccines.

Irilone, a Red Clover Isoflavone, Combined with Progesterone Enhances PR Signaling through the Estrogen and Glucocorticoid Receptors.

Trifolium pratense L. (red clover) is a popular botanical supplement used for women's health. Irilone isolated from red clover previously demonstrated progestogenic potentiation activity. In this study, irilone enhanced progesterone signaling was determined to not occur due to post-translational phosphorylation or by reducing progesterone receptor (PR) protein levels but instead increased PR protein levels in T47D breast cancer cells, which could be blocked by estrogen receptor (ER) antagonists, suggesting an ER dependent effect. Further, irilone increased luciferase activity from a hormone responsive element in a cell line that lacked ER and PR but expressed the glucocorticoid receptor (GR). A siRNA knockdown of GR in Ishikawa PR-B endometrial cancer cells reduced irilone's ability to enhance progesterone signaling. In an ovariectomized CD-1 mouse model, irilone did not induce uterine epithelial cell proliferation. The mechanism of action of irilone gives insight into PR crosstalk with other steroid hormone receptors, which can be important for understanding botanicals that are used for women's health.

Bioconjugate Supramolecular Pd2+ Metallacages Penetrate the Blood Brain Barrier In Vitro and In Vivo.

The biomedical application of discrete supramolecular metal-based structures, specifically self-assembled metallacages, is still an emergent field of study. Capitalizing on the knowledge gained in recent years on the development of 3-dimensional (3D) metallacages as novel drug delivery systems and theranostic agents, we explore here the possibility to target [Pd2L4]4+ cages (L = 3,5-bis(3-ethynylpyridine)phenyl ligand) to the brain. In detail, a new water-soluble homoleptic cage (CPepH3) tethered to a blood brain barrier (BBB)-translocating peptide was synthesized by a combination of solid-phase peptide synthesis (SPPS) and self-assembly procedures. The cage translocation efficacy was assessed by inductively coupled mass spectrometry (ICP-MS) in a BBB cellular model in vitro. Biodistribution studies of the radiolabeled cage [[99mTcO4]- ⊂ CPepH3] in the CD1 mice model demonstrate its brain penetration properties in vivo. Further DFT studies were conducted to model the structure of the [[99mTcO4]- ⊂ cage] complex. Moreover, the encapsulation capabilities and stability of the cage were investigated using the [ReO4]- anion, the "cold" analogue of [99mTcO4]-, by 1H NMR spectroscopy. Overall, our study constitutes another proof-of-concept of the unique potential of supramolecular coordination complexes for modifying the physiochemical and biodistribution properties of diagnostic species.

Role of Group 1 CD1-Restricted T Cells in Host Defense and Inflammatory Diseases.

Group 1 CD1-restricted T cells are members of the unconventional T cell family that recognize lipid antigens presented by CD1a, CD1b, and CD1c molecules. Although they developmentally mirror invariant natural killer T cells, they have diverse antigen specificity and functional capacity, with both anti-microbial and autoreactive targets. The role of group 1 CD1-restricted T cells has been best established in Mycobacterium tuberculosis (Mtb) infection in which a wide variety of lipid antigens have been identified and their ability to confer protection against Mtb infection in a CD1 transgenic mouse model has been shown. Group 1 CD1-restricted T cells have also been implicated in other infections, inflammatory conditions, and malignancies. In particular, autoreactive group 1 CD1-restricted T cells have been shown to play a role in several skin inflammatory conditions. The prevalence of group 1 CD1 autoreactive T cells in healthy individuals suggests the presence of regulatory mechanisms to suppress autoreactivity in homeostasis. The more recent use of group 1 CD1 tetramers and mouse models has allowed for better characterization of their phenotype, functional capacity, and underlying mechanisms of antigen-specific and autoreactive activation. These discoveries may pave the way for the development of novel vaccines and immunotherapies that target group 1 CD1-restricted T cells.

A Novel Outbred Mouse Model to Study Lung Memory Immunological Response Induced by Pertussis Vaccines.

Lung tissue resident memory (TRM) T cells can provide rapid and effective protective immunity against respiratory pathogens such as Bordetella pertussis We assessed an outbred CD1 mouse model and i.m. immunization to study vaccine-induced immune memory, using pertussis vaccines as an example. The phenotypes of cells from the lungs of CD1 mice that had been primed with either i.m. whole-cell B. pertussis (wP), acellular B. pertussis (aP) vaccines or buffer (unvaccinated) and challenged with B. pertussis were determined using flow cytometry and immunohistology. We observed a rapid and high increase of CD4+T cells expressing TRM markers by flow cytometry, supported by immunohistology observations, in lungs from wP-immunized mice. Priming mice with wP vaccine induced a more potent CD4+ response in lungs following B. pertussis challenge than priming with aP vaccine, although both were less potent than that observed in primoinfected mice. We also observed for the first time, to our knowledge, that CD8+ and γδ+ TRM-like T cell responses were induced in lungs of wP-primed mice postinfection. This novel outbred CD1 mouse model with i.m. immunization that enabled us to study vaccine-induced B. pertussis-specific memory T cells in lungs could be useful for evaluating candidate parenteral vaccines against B. pertussis or others pulmonary pathogens.

Sodium Hydrogen Exchanger Regulatory Factor-1 (NHERF1) Regulates Fetal Membrane Inflammation.

The fetal inflammatory response, a key contributor of infection-associated preterm birth (PTB), is mediated by nuclear factor kappa B (NF-kB) activation. Na+/H+ exchanger regulatory factor-1 (NHERF1) is an adapter protein that can regulate intracellular signal transduction and thus influence NF-kB activation. Accordingly, NHERF1 has been reported to enhance proinflammatory cytokine release and amplify inflammation in a NF-kB-dependent fashion in different cell types. The objective of this study was to examine the role of NHERF1 in regulating fetal membrane inflammation during PTB. We evaluated the levels of NHERF1 in human fetal membranes from term labor (TL), term not in labor (TNIL), and PTB and in a CD1 mouse model of PTB induced by lipopolysaccharide (LPS). Additionally, primary cultures of fetal membrane cells were treated with LPS, and NHERF1 expression and cytokine production were evaluated. Gene silencing methods using small interfering RNA targeting NHERF1 were used to determine the functional relevance of NHERF1 in primary cultures. NHERF1 expression was significantly (p < 0.001) higher in TL and PTB membranes compared to TNIL membranes, and this coincided with enhanced (p < 0.01) interleukin (IL)-6 and IL-8 expression levels. LPS-treated animals delivering PTB had increased levels of NHERF1, IL-6, and IL-8 compared to phosphate-buffered saline (PBS; control) animals. Silencing of NHERF1 expression resulted in a significant reduction in NF-kB activation and IL-6 and IL-8 production as well as increased IL-10 production. In conclusion, downregulation of NHERF1 increased anti-inflammatory IL-10, and reducing NHERF1 expression could be a potential therapeutic strategy to reduce the risk of infection/inflammation associated with PTB.

Growth Optimisation of Bacillus subtilis in medium supplemented with prebiotic gum Arabic

Bacillus subtilis is a type of gastrointestinal bacteria that build up a healthy gut microbiota. The bacterial species has been well documented to involve in maintaining a healthy gut homeostasis in which disruption in it’s microbial composition or “dysbiosis” has recently been linked to obesity. Probiotic in vivo restoration of dysbiosed microbiota have been proved successful in obese CD-1 mice model. However, prebiotic study using Gum Arabic (GA) assessing to optimise an in vitro lumen system specifically targeting to enhance the growth of B. subtilis are still lacking since this bacteria are depleted in obese individuals. Thus, this study aimed to establish the optimal growth conditions in simulated in vitro lumen system and to the best of our knowledge, this is the first in vitro study attempted to optimize the growth of B. subtilis in medium supplemented with prebiotic GA. Growth screening analysis suggested an optimal dosage of 1.0% and 0.5% glucose and GA, respectively. The highest growth rate was recorded at 0.7995 hours−1 with doubling time of 52.02 minutes with extended period of stationary phase. The optimal GA concentration and fermentation conditions were determined at 0.67%, pH 7.4 and temperature of 37°C. The validated suggested model indicates that the supplementation of GA into an optimal fermentation systems is promising to enhance the growth of gut microflora Bacillus subtilis, for restoration of a dysbiosed gut microbiota in vitro.

P0679LOSARTAN REDUCES INFLAMMATION INDUCED BY TERTIARY LYMPHOID TISSUES IN ENSUING CKD AFTER ACUTE KIDNEY INJURY

Abstract Background and Aims Chronic inflammation and renin-angiotensin system (RAS) overactivation have been demonstrated as a central mechanism that results in the development of progressive chronic kidney disease (CKD). Evidences have indicated that tertiary lymphoid tissues (TLT) form after acute kidney injury (AKI), especially in old-aged mice. Heterogeneous fibroblasts which appear and secret proinflammatory cytokines after AKI underlie TLT formation. There is an association between TLT sizes and impaired renal function along with increase of proinflammatory cytokines, suggesting a probable cause of persistent inflammation after AKI. In this study, we explored the relationship between TLT development and RAS activity. The phenotypes of heterogeneous fibroblasts diversified after AKI is also investigated. Method Here we utilized ischemia-reperfusion injury to cause ensuing CKD both in a CD-1 and C57BL6J mice model. We also used staining of CD3 and B220 to label and confirm multiple TLTs which represents T and B lymphocytes respectively. Results Administration of losartan, a kind of RAS inhibitor, offered protection against subsequent CKD development and attenuated number and size of TLT besides blood pressure reduction. Moreover, heterogeneous fibroblasts diminished and proinflammatory cytokines such as CXCL13, CCL21 and CCL19 which trigger the initiation of TLT were downregulated. In experiments of several fibroblast cell lines, angiotensin II (Ang II) administration could induce differentiation of fibroblasts with distinct phenotypes expressing p75 neurotrophin receptor and retinaldehyde dehydrogenase 2 which contribute to TLT formation. These fibroblasts expressing PDGFRβ stimulated production of aforementioned proinflammatory cytokines and expression of pro-fibrotic genes. Conclusion Our data demonstrates that TLT plays an important role in AKI-CKD transition. We also verify that Ang II causes phenotypic change of fibroblasts which constitute TLT. Losartan can ameliorate TLT formation through inhibition of these distinctive fibroblasts. It is a plausible mechanism to reduce ensuing CKD.

Dextran sulfate sodium mouse model of inflammatory bowel disease evaluated for systemic genotoxicity via blood micronucleus and Pig-a gene mutation assays

Inflammatory bowel disease (IBD) is an important risk factor for gastrointestinal cancers. Inflammation and other carcinogenesis-related effects at distal, tissue-specific sites require further study. In order to better understand if systemic genotoxicity is associated with IBD, we exposed mice to dextran sulfate sodium salt (DSS) and measured the incidence of micronucleated cells (MN) and Pig-a mutant phenotype cells in blood erythrocyte populations. In one study, 8-week-old male CD-1 mice were exposed to 0, 1, 2, 3 or 4% w/v DSS in drinking water. The 4-week in-life period was divided into four 1-week intervals-alternately on then off DSS treatment. Low volume blood samples were collected for MN analysis at the end of each week, and cardiac blood samples were collected at the end of the 4-week period for Pig-a analyses. The two highest doses of DSS were observed to induce significant increases in reticulocyte frequencies. Even so, no statistically significant treatment-related effects on the genotoxicity biomarkers were evident. While one high-dose mouse showed modestly elevated MN frequencies during the DSS treatment cycles, it also exhibited exceptionally high reticulocyte frequencies (e.g. 18.7% at the end of the second DSS cycle). In a second study, mice were treated with 0 or 4% DSS for 9-18 consecutive days. Exposure was continued until rectal bleeding or morbidity was evident, at which point the treatment was terminated and blood was collected for MN analysis. The Pig-a assay was conducted on samples collected 29 days after the start of treatment. The initial blood specimens showed highly elevated reticulocyte frequencies in DSS-exposed mice (mean ± SEM = 1.75 ± 0.10% vs. 13.04 ± 3.66% for 0 vs. 4% mice, respectively). Statistical analyses showed no treatment-related effect on MN or Pig-a mutant frequencies. Even so, the incidence of MN versus reticulocytes in the DSS-exposed mice were positively correlated (linear fit R2 = 0.657, P = 0.0044). Collectively, these results suggest that in the case of the DSS CD-1 mouse model, systemic effects include stress erythropoiesis but not remarkable genotoxicity. To the extent MN may have been slightly elevated in a minority of individual mice, these effects appear to be secondary, likely attributable to stimulated erythropoiesis.

Gold Nanoparticles Crossing Blood-Brain Barrier Prevent HSV-1 Infection and Reduce Herpes Associated Amyloid-βsecretion.

Infections caused by HSV-1 and their typical outbreaks invading the nervous system have been related to neurodegenerative diseases. HSV-1 infection may deregulate the balance between the amyloidogenic and non-amyloidogenic pathways, raising the accumulation of amyloid-β peptides, one of the hallmarks in the neurodegenerative diseases. An effective treatment against both, HSV-1 infections and neurodegeneration, is a major therapeutic target. Therefore, gold nanoparticles (NPAus) have been previously studied in immunotherapy, cancer and cellular disruptions with very promising results. Our study demonstrates that a new NPAus family inhibits the HSV-1 infection in a neural-derived SK-N-MC cell line model and that this new NPAus reduces the HSV-1-induced β-secretase activity, as well as amyloid-β accumulation in SK-APP-D1 modifies cell line. We demonstrated that NPAuG3-S8 crosses the blood-brain barrier (BBB) and does not generate cerebral damage to in vivo CD1 mice model. The NPAuG3-S8 could be a promising treatment against neuronal HSV-1 infections and neuronal disorders related to the Aβ peptides.

Zika structural genes determine the virulence of African and Asian lineages

ABSTRACT The Asian lineage of Zika virus (ZIKV) is responsible for the recent epidemics in the Americas and severe disease, whereas the African lineage of ZIKV has not been reported to cause epidemics or severe disease. We constructed a cDNA infectious clone (IC) of an African ZIKV strain, which, together with our previously developed Asian ZIKV strain IC, allowed us to engineer chimeric viruses by swapping the structural and non-structural genes between the two lineages. Recombinant parental and chimeric viruses were analyzed in A129 and newborn CD1 mouse models. In the A129 mice, the African strain developed higher viremia, organ viral loading, and mortality rate. In CD1 mice, the African strain exhibited a higher neurovirulence than the Asian strain. A chimeric virus containing the structural genes from the African strain is more virulent than the Asian strain, whereas a chimeric virus containing the non-structural genes from the African strain exhibited a virulence comparable to the Asian strain. These results suggest that (i) African strain is more virulent than Asian strain and (ii) viral structural genes primarily determine the virulence difference between the two lineages in mouse models. Other factors may contribute to the discrepancy between the mouse and epidemic results.

Rescue of PB-resistant neonatal seizures with single-dose of small-molecule TrkB antagonist show long-term benefits

A recently characterized CD-1 mouse model of phenobarbital (PB)-resistant neonatal ischemic-seizures (i.e.; unilateral carotid ligation) was shown to be associated with age-dependent (P7 vs. P10) acute seizure severity and PB-efficacy (i.e.; PB-resistant vs. PB-responsive). ANA12, a novel small-molecule TrkB antagonist, rescued the PB-resistance at P7 in a dose-dependent manner and prevented the post-ischemic downregulation of KCC2, the chief Cl- extruder in neurons. The long-term consequences of this novel rescue-intervention with ANA12 + PB in P7 and P10 ligated pups was investigated and compared to the standard first-line protocol of PB-alone loading dose. The mice underwent neurobehavioral testing, 24 h video-EEG-EMG monitoring, and immunohistochemistry in ipsi- and contralateral cortices as adults following the neonatal interventions. ANA12 + PB rescued the emergence of hyperactivity in post-ischemic P7, but not in P10 pups as adults. ANA12 + PB administration at neither P7 nor P10 significantly altered 24 h macro-sleep architecture in adults when compared to PB-alone. Behavioral state-dependent gamma (35−50 Hz) power homeostasis showed the most significant between-group differences that were age-dependent. ANA12 + PB treatment, but not PB-alone, rescued the loss of gamma power homeostasis present in P7 ligate-control but absent in P10 ligate group, highlighting the age-dependence. In contrast, PB-alone treatment, but not ANA12+PB, significantly reduced the elevated delta-AUC observed in P10 ligate-controls, when PB is efficacious by itself. These results indicate that the rescue of acute PB-resistant neonatal seizures using a novel intervention positively modulates the long-term outcomes at P7 when the seizures are refractory.