Role of cancer stem cell ecosystem on breast cancer metastasis and related mouse models

Breast cancer metastasis is responsible for most breast cancer-related deaths and is influenced by many factors within the tumor ecosystem,including tumor cells and microenvironment.Breast cancer stem cells(BCSCs)constitute a small population of cancer cells with unique characteristics,including their capacity for self-renewal and differentiation.Studies have shown that BCSCs not only drive tumorigenesis but also play a crucial role in promoting metastasis in breast cancer.The tumor microenvironment(TME),composed of stromal cells,immune cells,blood vessel cells,fibroblasts,and microbes in proximity to cancer cells,is increasingly recognized for its crosstalk with BCSCs and role in BCSC survival,growth,and dissemination,thereby influencing metastatic ability.Hence,a thorough understanding of BCSCs and the TME is critical for unraveling the mechanisms underlying breast cancer metastasis.In this review,we summarize current knowledge on the roles of BCSCs and the TME in breast cancer metastasis,as well as the underlying regulatory mechanisms.Furthermore,we provide an overview of relevant mouse models used to study breast cancer metastasis,as well as treatment strategies and clinical trials addressing BCSC-TME interactions during metastasis.Overall,this study provides valuable insights for the development of effective therapeutic strategies to reduce breast cancer metastasis.

Inhibition of Bcl-6 Expression Ameliorates Asthmatic Characteristics in Mice

Objective The function of Bcl-6 in T follicular helper(Tfh)cell maturation is indispensable,and Tfh cells play a pivotal role in asthma.This study investigated the impact of Bcl-6 on asthmatic traits.Methods The microscopic pathological alterations,airway resistance(AR),and lung compliance(LC)were determined in asthmatic mice and Bcl-6 interference mice.The surface molecular markers of Tfh cells and the Bcl-6 mRNA and protein expression were determined by flow cytometry,RT-qPCR,and Western blotting,respectively.The relationships between the Tfh cell ratio and the IgE and IgG1 concentrations in peripheral blood mononuclear cells(PBMCs)and bronchoalveolar lavage fluid(BALF)were determined.Results Asthmatic inflammatory changes were observed in the lung tissue and were attenuated by Bcl-6 siRNA and dexamethasone(DXM).Asthmatic mice exhibited an increased AR and a decreased LC,while Bcl-6 siRNA or DXM mitigated these changes.The percentages of Tfh cells and eosinophils were significantly increased in the asthmatic mice,and they significantly decreased after Bcl-6 inhibition or DXM treatment.RT-qPCR and Western blotting analyses revealed that the Bcl-6 expression level in PBMCs was significantly higher in asthmatic mice,and it decreased following Bcl-6 inhibition or DXM treatment.The IgE expression in the serum and BALF and the B cell expression in PBMCs exhibited a similar trend.In asthmatic mice,the ratio of Tfh cells in the peripheral blood showed a strong positive correlation with the IgE levels in the serum and BALF,but not with the IgG1 levels.Conclusion The amelioration of airway inflammation and airway hyper-responsiveness is achieved through Bcl-6 suppression,which effectively hinders Tfh cell differentiation,ultimately resulting in a concurrent reduction in IgE production.

Simple procedure for assessing diffuse subarachnoid hemorrhage successfully created using filament perforation method in mice

The murine model of subarachnoid hemorrhage(SAH)is a valuable experimental tool for investigating molecular and cellular mechanisms,and the endovascular filament perforation technique can be used to simulate prominent pathophysiological features observed after human SAH;however,current validation methods for assessing an appropriate SAH model are limited.Here,we introduce a simple procedure for se-lecting a mouse model of diffuse SAH.SAH was induced in 24 mice using a standard filament perforation method.After confirming survival at 24 h,SAH was scored 0-1 based on T2*-weighted images on whole-brain magnetic resonance imaging(MRI)and visual surveillance of the cisterna magna(CM)through the dura mater.The CM-based SAH grading correlated well with a reference parameter defined by extracted brain(r^(2)=0.53,p<0.0001).The receiver operating characteristic curve revealed a sensi-tivity of 85%and a specificity of 91%for detecting diffuse SAH,with a similar area under the curve(0.89±0.06[standard error of the mean])as the MRI-based grading(0.72±0.10,p=0.12).Our data suggest that confirming an SAH clot in the CM is a valuable way to select a clinically relevant diffuse SAH model that can be used in future experimental studies.

Comparison of immune responses and intestinal flora in epicutaneously sensitized BALB/c or C57BL/6 mouse models of food allergy

Cutaneous exposure to food allergens through a disrupted skin barrier is recognized as an important cause of food allergy,and the cutaneous sensitized mouse model has been established to investigate relevant allergic disorders.However,the role of different genetic backgrounds of mice on immune responses to food allergens upon epicutaneous sensitization is largely unknown.In this study,two strains of mice,i.e.,the BALB/c and C57BL/6 mice,were epicutaneously sensitized with ovalbumin on atopic dermatitis(AD)-like skin lesions,followed by intragastric challenge to induce IgE-mediated food allergy.Allergic outcomes were measured as clinical signs,specific antibodies and cytokines,and immune cell subpopulations,as well as changes in intestinal barrier function and gut microbiota.Results showed that both strains of mice exhibited typical food-allergic symptoms with a Th2-skewed response.The C57BL/6 mice,rather than the BALB/c mice,were fitter for establishing an epicutaneously sensitized model of food allergy since a stronger Th2-biased response and severer disruptions in the intestinal barrier and gut homeostasis were observed.This study provides knowledge for selecting an appropriate mouse model to study food-allergic responses associated with AD-like skin lesions and highlights the role of genetic variations in the immune mechanism underlying pathogenesis of food allergy.

Identifying Brand Consistency by Product Differentiation Using CNN

This paper presents a new method of using a convolutional neural network(CNN)in machine learning to identify brand consistency by product appearance variation.In Experiment 1,we collected fifty mouse devices from the past thirty-five years from a renowned company to build a dataset consisting of product pictures with pre-defined design features of their appearance and functions.Results show that it is a challenge to distinguish periods for the subtle evolution of themouse devices with such traditionalmethods as time series analysis and principal component analysis(PCA).In Experiment 2,we applied deep learning to predict the extent to which the product appearance variation ofmouse devices of various brands.The investigation collected 6,042 images ofmouse devices and divided theminto the Early Stage and the Late Stage.Results show the highest accuracy of 81.4%with the CNNmodel,and the evaluation score of brand style consistency is 0.36,implying that the brand consistency score converted by the CNN accuracy rate is not always perfect in the real world.The relationship between product appearance variation,brand style consistency,and evaluation score is beneficial for predicting new product styles and future product style roadmaps.In addition,the CNN heat maps highlight the critical areas of design features of different styles,providing alternative clues related to the blurred boundary.The study provides insights into practical problems for designers,manufacturers,and marketers in product design.It not only contributes to the scientific understanding of design development,but also provides industry professionals with practical tools and methods to improve the design process and maintain brand consistency.Designers can use these techniques to find features that influence brand style.Then,capture these features as innovative design elements and maintain core brand values.

Smad8 is involvement in follicular development via the regulation of granulosa cell growth and steroidogenesis in mice

Background:SMAD family proteins(SMADs)are crucial transcription factors downstream of transforming growth factor beta(TGF-ß)/SMAD signaling pathways that have been reported to play a pivotal role in mammalian reproduction.However,the role of SMAD family member 8(SMAD8,also known as SMAD9),a member of the SMAD family,in mammalian reproduction remains unclear.Methods:We employed RNA interference techniques to knock down Smad8 expression in mouse granulosa cells(GCs)to investigate the effects of Smad8 on GC growth and steroidogenesis.Results:Our findings revealed a significant decrease in the proliferative capacity and a substantial increase in the apoptosis rate of GCs after transfection with Smad8-siRNA for 48 h.Subsequent hormone assays demonstrated a significant decrease in estradiol(E2)levels,whereas progesterone(P4)remained unchanged.Further mechanistic analysis showed that the mRNA expression of proliferating cell nuclear antigen(Pcna),Cyclin D2,cell cycle-dependent kinase 4(Cdk4),B-cell lymphoma-2(Bcl-2),estrogen receptor(Er),luteinizing hormone receptor(Lhr)and cytochrome P450 family 19 subfamily A member 1(Cyp19a1)significantly decreased.Conversely,the mRNA of cysteine aspartate proteinase 3(Caspase 3)significantly increased,wheras Bcl2-associated X(Bax),folliclestimulating hormone receptor(Fshr)and cytochrome P450 family 11 subfamily A member 1(Cyp11a1)remained unchanged compared to the controls.Conclusion:This study indicates that Smad8 knockdown inhibits cell proliferation,promotes apoptosis,reduces Er and Lhr transcription,and decreases E2 production in mouse GCs.These findings suggest that Smad8 may serve as a novel genetic marker for mammalian reproduction.

Magnesium-L-threonate treats Alzheimer's disease by modulating the microbiota-gut-brain axis

Disturbances in the microbiota-gut-brain axis may contribute to the development of Alzheimer's disease. Magnesium-L-threonate has recently been found to have protective effects on learning and memory in aged and Alzheimer's disease model mice. However, the effects of magnesium-L-threonate on the gut microbiota in Alzheimer's disease remain unknown. Previously, we reported that magnesium-L-threonate treatment improved cognition and reduced oxidative stress and inflammation in a double-transgenic line of Alzheimer's disease model mice expressing the amyloid-β precursor protein and mutant human presenilin 1(APP/PS1). Here, we performed 16S r RNA amplicon sequencing and liquid chromatography-mass spectrometry to analyze changes in the microbiome and serum metabolome following magnesium-Lthreonate exposure in a similar mouse model. Magnesium-L-threonate modulated the abundance of three genera in the gut microbiota, decreasing Allobaculum and increasing Bifidobacterium and Turicibacter. We also found that differential metabolites in the magnesiumL-threonate-regulated serum were enriched in various pathways associated with neurodegenerative diseases. The western blotting detection on intestinal tight junction proteins(zona occludens 1, occludin, and claudin-5) showed that magnesium-L-threonate repaired the intestinal barrier dysfunction of APP/PS1 mice. These findings suggest that magnesium-L-threonate may reduce the clinical manifestations of Alzheimer's disease through the microbiota-gut-brain axis in model mice, providing an experimental basis for the clinical treatment of Alzheimer's disease.

Skeletal muscle as a molecular and cellular biomarker of disease progression in amyotrophic lateral sclerosis:a narrative review

Amyotrophic lateral sclerosis is a fatal multisystemic neurodegenerative disease with motor neurons being a primary target.Although progressive weakness is a hallmark feature of amyotrophic lateral sclerosis,there is considerable heterogeneity,including clinical presentation,progression,and the underlying triggers for disease initiation.Based on longitudinal studies with families harboring amyotrophic lateral sclerosis-associated gene mutations,it has become apparent that overt disease is preceded by a prodromal phase,possibly in years,where compensatory mechanisms delay symptom onset.Since 85-90%of amyotrophic lateral sclerosis is sporadic,there is a strong need for identifying biomarkers that can detect this prodromal phase as motor neurons have limited capacity for regeneration.Current Food and Drug Administration-approved therapies work by slowing the degenerative process and are most effective early in the disease.Skeletal muscle,including the neuromuscular junction,manifests abnormalities at the earliest stages of the disease,before motor neuron loss,making it a promising source for identifying biomarkers of the prodromal phase.The accessibility of muscle through biopsy provides a lens into the distal motor system at earlier stages and in real time.The advent of“omics”technology has led to the identification of numerous dysregulated molecules in amyotrophic lateral sclerosis muscle,ranging from coding and non-coding RNAs to proteins and metabolites.This technology has opened the door for identifying biomarkers of disease activity and providing insight into disease mechanisms.A major challenge is correlating the myriad of dysregulated molecules with clinical or histological progression and understanding their relevance to presymptomatic phases of disease.There are two major goals of this review.The first is to summarize some of the biomarkers identified in human amyotrophic lateral sclerosis muscle that have a clinicopathological correlation with disease activity,evidence of a similar dysregulation in the SOD1G93A mouse during presymptomatic stages,and evidence of progressive change during disease progression.The second goal is to review the molecular pathways these biomarkers reflect and their potential role in mitigating or promoting disease progression,and as such,their potential as therapeutic targets in amyotrophic lateral sclerosis.

Cardiac differentiation is modulated by anti-apoptotic signals in murine embryonic stem cells

BACKGROUND Embryonic stem cells(ESCs)serve as a crucial ex vivo model,representing epiblast cells derived from the inner cell mass of blastocyst-stage embryos.ESCs exhibit a unique combination of self-renewal potency,unlimited proliferation,and pluripotency.The latter is evident by the ability of the isolated cells to differ-entiate spontaneously into multiple cell lineages,representing the three primary embryonic germ layers.Multiple regulatory networks guide ESCs,directing their self-renewal and lineage-specific differentiation.Apoptosis,or programmed cell death,emerges as a key event involved in sculpting and forming various organs and structures ensuring proper embryonic development.How-ever,the molecular mechanisms underlying the dynamic interplay between diffe-rentiation and apoptosis remain poorly understood.AIM To investigate the regulatory impact of apoptosis on the early differentiation of ESCs into cardiac cells,using mouse ESC(mESC)models-mESC-B-cell lym-phoma 2(BCL-2),mESC-PIM-2,and mESC-metallothionein-1(MET-1)-which overexpress the anti-apoptotic genes Bcl-2,Pim-2,and Met-1,respectively.METHODS mESC-T2(wild-type),mESC-BCL-2,mESC-PIM-2,and mESC-MET-1 have been used to assess the effect of potentiated apoptotic signals on cardiac differentiation.The hanging drop method was adopted to generate embryoid bodies(EBs)and induce terminal differentiation of mESCs.The size of the generated EBs was measured in each condition compared to the wild type.At the functional level,the percentage of cardiac differentiation was measured by calculating the number of beating cardiomyocytes in the manipulated mESCs compared to the control.At the molecular level,quantitative reverse transcription-polymerase chain reaction was used to assess the mRNA expression of three cardiac markers:Troponin T,GATA4,and NKX2.5.Additionally,troponin T protein expression was evaluated through immunofluorescence and western blot assays.RESULTS Our findings showed that the upregulation of Bcl-2,Pim-2,and Met-1 genes led to a reduction in the size of the EBs derived from the manipulated mESCs,in comparison with their wild-type counterpart.Additionally,a decrease in the count of beating cardiomyocytes among differentiated cells was observed.Furthermore,the mRNA expression of three cardiac markers-troponin T,GATA4,and NKX2.5-was diminished in mESCs overexpressing the three anti-apoptotic genes compared to the control cell line.Moreover,the overexpression of the anti-apoptotic genes resulted in a reduction in troponin T protein expression.CONCLUSION Our findings revealed that the upregulation of Bcl-2,Pim-2,and Met-1 genes altered cardiac differentiation,providing insight into the intricate interplay between apoptosis and ESC fate determination.

MicroRNA-630 alleviates inflammatory reactions in rats with diabetic kidney disease by targeting toll-like receptor 4

BACKGROUND Diabetic kidney disease(DKD)is a major complication of diabetes mellitus.Renal tubular epithelial cell(TEC)damage,which is strongly associated with the inflammatory response and mesenchymal trans-differentiation,plays a significant role in DKD;However,the precise molecular mechanism is unknown.The recently identified microRNA-630(miR-630)has been hypothesized to be closely associated with cell migration,apoptosis,and autophagy.However,the association between miR-630 and DKD and the underlying mechanism remain unknown.AIM To investigate how miR-630 affects TEC injury and the inflammatory response in DKD rats.METHODS Streptozotocin was administered to six-week-old male rats to create a hypergly cemic diabetic model.In the second week of modeling,the rats were divided into control,DKD,negative control of lentivirus,and miR-630 overexpression groups.After 8 wk,urine and blood samples were collected for the kidney injury assays,and renal tissues were removed for further molecular assays.The target gene for miR-630 was predicted using bioinformatics,and the association between miR-630 and toll-like receptor 4(TLR4)was confirmed using in vitro investigations and double luciferase reporter gene assays.Overexpression of miR-630 in DKD rats led to changes in body weight,renal weight index,basic blood parameters and histopathological changes.RESULTS The expression level of miR-630 was reduced in the kidney tissue of rats with DKD(P<0.05).The miR-630 and TLR4 expressions in rat renal TECs(NRK-52E)were measured using quantitative reverse transcription polymerase chain reaction.The mRNA expression level of miR-630 was significantly lower in the high-glucose(HG)and HG+mimic negative control(NC)groups than in the normal glucose(NG)group(P<0.05).In contrast,the mRNA expression level of TLR4 was significantly higher in these groups(P<0.05).However,miR-630 mRNA expression increased and TLR4 mRNA expression significantly decreased in the HG+miR-630 mimic group than in the HG+mimic NC group(P<0.05).Furthermore,the levels of tumor necrosis factor-alpha(TNF-α),interleukin-1β(IL-1β),and IL-6 were significantly higher in the HG and HG+mimic NC groups than in NG group(P<0.05).However,the levels of these cytokines were significantly lower in the HG+miR-630 mimic group than in the HG+mimic NC group(P<0.05).Notably,changes in protein expression were observed.The HG and HG+mimic NC groups showed a significant decrease in E-cadherin protein expression,whereas TLR4,α-smooth muscle actin(SMA),and collagen IV protein expression increased(P<0.05).Conversely,the HG+miR-630 mimic group exhibited a significant increase in E-cadherin protein expression and a notable decrease in TLR4,α-SMA,and collagen IV protein expression than in the HG+mimic NC group(P<0.05).The miR-630 targets TLR4 gene expression.In vivo experiments demonstrated that DKD rats treated with miR-630 agomir exhibited significantly higher miR-630 mRNA expression than DKD rats injected with agomir NC.Additionally,rats treated with miR-630 agomir showed significant reductions in urinary albumin,blood glucose,TLR4,and proinflammatory markers(TNF-α,IL-1β,and IL-6)expression levels(P<0.05).Moreover,these rats exhibited fewer kidney lesions and reduced infiltration of inflammatory cells.CONCLUSION MiR-630 may inhibit the inflammatory reaction of DKD by targeting TLR4,and has a protective effect on DKD.

Expression levels of K_(ATP)channel subunits and morphological changes in the mouse liver after exposure to radiation

BACKGROUND ATP sensitive K+(K_(ATP))channels are ubiquitously distributed in various of cells and tissues,including the liver.They play a role in the pathogenesis of myocardial and liver ischemia.AIM To evaluate the radiation-induced changes in the expression of K_(ATP)channel subunits in the mouse liver to understand the potential role of K_(ATP)channels in radiation injury.METHODS Adult C57BL/6 mice were randomly exposed toγ-rays at 0 Gy(control,n=2),0.2 Gy(n=6),1 Gy(n=6),or 5 Gy(n=6).The livers were removed 3 and 24 h after radiation exposure.Hematoxylin and eosin staining was used for morphological observation;immunohistochemical staining was applied to determine the expression of K_(ATP)channel subunits in the liver tissue.RESULTS Compared with the control group,the livers exposed to 0.2 Gyγ-ray showed an initial increase in the expression of Kir6.1 at 3 h,followed by recovery at 24 h after exposure.Exposure to a high dose of 5.0 Gy resulted in decreased expression of Kir6.1 and increased expression of SUR2B at 24 h.However,the expression of Kir6.2,SUR1,or SUR2A had no remarkable changes at 3 and 24 h after exposure to any of these doses.CONCLUSION The expression levels of Kir6.1 and SUR2B in mouse liver changed differently in response to different radiation doses,suggesting a potential role for them in radiation-induced liver injury.

小鼠整胚石蜡切片制备方法的改进

Objective:The morphology analysis of whole-mount mouse embryos was observed using an improved paraffin section technique.Methods:Mouse embryos of varying embryonic ages were collected and whole-mount embryo paraffin sections were prepared using PFA-intravenously injected fixation,prolonged dehydration,and paraffin embedding.Hematoxylin and eosin(H&E)staining and immunohistochemical staining were employed to evaluate the quality of sections,with different tissues being observed labeled by CD34.Results:Following a series of tissue processing and staining procedures,the structure of the whole-mount mouse embryo was well-preserved,and the staining was clear and easily distinguishable.Embryos of different embryonic ages were treated differently,yet the quality of tissue processing remained highly consistent.Conclusion:Tissue processing and staining have been significantly improved,allowing for the easy acquisition of whole-mount mouse embryos of different ages through simplified methods of tissue fixation and dehydration duration.The staining results are clear and stable,providing technical support for the study of mouse embryo development.

MicroRNAs in mouse and rat models of experimental epilepsy and potential therapeutic targets

Epilepsy is a common and serious neurological disease that causes recurrent seizures. The brain damage caused by seizures can lead to depression, anxiety, cognitive impairment, or disability. In almost all cases chronic seizures are difficult to cure. MicroRNAs are widely expressed in the central nervous system and play important roles in the pathogenesis of several neurological disorders, including epilepsy. A variety of animals(mostly mice and rats) have been used to induce experimental epilepsy using different protocols and miRNA profiling performed. Most of the recent studies reviewed had performed miRNA profiling in hippocampal tissues and a large number of microRNAs were dysregulated when compared to controls. Most notably, miR-132-3p,-146a-5p,-10a-5p,-21a-3p,-27a-3p,-142a-5p,-212-3p,-431-5p, and-155 were upregulated in both the mouse and rat studies. Overexpression of miR-137 and miR-219 decreased seizure severity in a mouse epileptic model, and suppression of miR-451,-10a-5p,-21a-5p,-27a-5p,-142a-5p,-431-5p,-155, and-134 had a positive influence on seizure behavior. In the rat studies, overexpression of miR-139-5p decreased neuronal damage in drug-resistant rats and inhibition of miR-129-2-3p,-27a-3p,-155,-134,-181a, and-146a had a positive effect on seizure behavior and/or reduced the loss of neuronal cells. Further studies are warranted using adult female and immature male and female animals. It would also be helpful to test the ability of specific agomirs and antagomirs to control seizure activity in a subhuman primate model of epilepsy such as adult marmosets injected intraperitoneally with pilocarpine or cynomolgus monkeys given intrahippocampal injections of kainic acid.

High-fat diet and oral infection induced type 2 diabetes and obesity development under different genetic backgrounds

Background:Type 2 diabetes(T2D)is an adult-onset and obese form of diabetes caused by an interplay between genetic,epigenetic,and environmental components.Here,we have assessed a cohort of 11 genetically different collaborative cross(CC)mouse lines comprised of both sexes for T2D and obesity developments in response to oral infection and high-fat diet(HFD)challenges.Methods:Mice were fed with either the HFD or the standard chow diet(control group)for 12 weeks starting at the age of 8 weeks.At week 5 of the experiment,half of the mice of each diet group were infected with Porphyromonas gingivalis and Fusobacterium nucleatum bacteria strains.Throughout the 12-week experimental period,body weight(BW)was recorded biweekly,and intraperitoneal glucose tolerance tests were performed at weeks 6 and 12 of the experiment to evaluate the glucose tolerance status of mice.Results:Statistical analysis has shown the significance of phenotypic variations between the CC lines,which have different genetic backgrounds and sex effects in different experimental groups.The heritability of the studied phenotypes was estimated and ranged between 0.45 and 0.85.We applied machine learning methods to make an early call for T2D and its prognosis.The results showed that classification with random forest could reach the highest accuracy classification(ACC=0.91)when all the attributes were used.Conclusion:Using sex,diet,infection status,initial BW,and area under the curve(AUC)at week 6,we could classify the final phenotypes/outcomes at the end stage of the experiment(at 12 weeks).

CMT1A current gene therapy approaches and promising biomarkers

Charcot-Marie-Tooth neuropathies(CMT)constitute a group of common but highly heterogeneous,non-syndromic genetic disorders affecting predominantly the peripheral nervous system.CMT type 1A(CMT1A)is the most frequent type and accounts for almost~50%of all diagnosed CMT cases.CMT1A results from the duplication of the peripheral myelin protein 22(PMP22)gene.Overexpression of PMP22 protein overloads the protein folding apparatus in Schwann cells and activates the unfolded protein response.This leads to Schwann cell apoptosis,dys-and de-myelination and secondary axonal degeneration,ultimately causing neurological disabilities.During the last decades,several different gene therapies have been developed to treat CMT1A.Almost all of them remain at the pre-clinical stage using CMT1A animal models overexpressing PMP22.The therapeutic goal is to achieve gene silencing,directly or indirectly,thereby reversing the CMT1A genetic mechanism allowing the recovery of myelination and prevention of axonal loss.As promising treatments are rapidly emerging,treatment-responsive and clinically relevant biomarkers are becoming necessary.These biomarkers and sensitive clinical evaluation tools will facilitate the design and successful completion of future clinical trials for CMT1A.

Anti-infection effects of heparin on SARS-CoV-2 in a diabetic mouse model

Severe acute respiratory syndrome coronavirus 2(SARSCo V-2)infection can result in more severe syndromes and poorer outcomes in patients with diabetes and obesity.However,the precise mechanisms responsible for the combined impact of coronavirus disease 2019(COVID-19)and diabetes have not yet been elucidated,and effective treatment options for SARS-Co V-2-infected diabetic patients remain limited.To investigate the disease pathogenesis,K18-h ACE2 transgenic(h ACE2^(Tg))mice with a leptin receptor deficiency(h ACE2-Lepr^(-/-))and high-fat diet(h ACE2-HFD)background were generated.The two mouse models were intranasally infected with a 5×10^(5) median tissue culture infectious dose(TCID_(50))of SARSCo V-2,with serum and lung tissue samples collected at 3days post-infection.The h ACE2-Lepr^(-/-)mice were then administered a combination of low-molecular-weight heparin(LMWH)(1 mg/kg or 5 mg/kg)and insulin via subcutaneous injection prior to intranasal infection with1×10^(4) TCID_(50)of SARS-Co V-2.Daily drug administration continued until the euthanasia of the mice.Analyses of viral RNA loads,histopathological changes in lung tissue,and inflammation factors were conducted.Results demonstrated similar SARS-Co V-2 susceptibility in h ACE2^(Tg)mice under both lean(chow diet)and obese(HFD)conditions.However,compared to the h ACE2-Lepr^(+/+)mice,h ACE2-Lepr^(-/-)mice exhibited more severe lung injury,enhanced expression of inflammatory cytokines and hypoxia-inducible factor-1α(HIF-1α),and increased apoptosis.Moreover,combined LMWH and insulin treatment effectively reduced disease progression and severity,attenuated lung pathological changes,and mitigated inflammatory responses.In conclusion,preexisting diabetes can lead to more severe lung damage upon SARS-Co V-2 infection,and LMWH may be a valuable therapeutic approach for managing COVID-19patients with diabetes.

Characteristics of traumatic brain injury models:from macroscopic blood flow changes to microscopic mitochondrial changes

Controlled cortical impingement is a widely accepted method to induce traumatic brain injury to establish a traumatic brain injury animal model.A strike depth of 1 mm at a certain speed is recommended for a moderate brain injury and a depth of>2 mm is used to induce severe brain injury.However,the different effects and underlying mechanisms of these two model types have not been proven.This study investigated the changes in cerebral blood flow,differences in the degree of cortical damage,and differences in motor function under different injury parameters of 1 and 2 mm at injury speeds of 3,4,and 5 m/s.We also explored the functional changes and mitochondrial damage between the 1 and 2 mm groups in the acute(7 days)and chronic phases(30 days).The results showed that the cerebral blood flow in the injured area of the 1 mm group was significantly increased,and swelling and bulging of brain tissue,increased vascular permeability,and large-scale exudation occurred.In the 2 mm group,the main pathological changes were decreased cerebral blood flow,brain tissue loss,and cerebral vasospasm occlusion in the injured area.Substantial motor and cognitive impairments were found on day 7 after injury in the 2 mm group;at 30 days after injury,the motor function of the 2 mm group mice recovered significantly while cognitive impairment persisted.Transcriptome sequencing showed that compared with the 1 mm group,the 2 mm group expressed more ferroptosis-related genes.Morphological changes of mitochondria in the two groups on days 7 and 30 using transmission electron microscopy revealed that on day 7,the mitochondria in both groups shrank and the vacuoles became larger;on day 30,the mitochondria in the 1 mm group became larger,and the vacuoles in the 2 mm group remained enlarged.By analyzing the proportion of mitochondrial subgroups in different groups,we found that the model mice had different patterns of mitochondrial composition at different time periods,suggesting that the difference in the degree of damage among traumatic brain injury groups may reflect the mitochondrial changes.Taken together,differences in mitochondrial morphology and function between the 1 and 2 mm groups provide a new direction for the accurate classification of traumatic brain injury.Our results provide reliable data support and evaluation methods for promoting the establishment of standard mouse controlled cortical impingement model guidelines.

Therapeutic effect of folic acid combined with decitabine on diabetic mice

AIM:To evaluate the therapeutic effect of folic acid combined with decitabine on diabetic mice.METHODS:The diabetic model of db/db mice were randomly divided into model group,folic acid group,decitabine group,folic acid combined with decitabine group,and C57 mice as normal control group.The density of retinal blood vessels and retinal thickness were detected by fundus photography and optical coherence tomography,respectively.Pathological changes of retina were observed by hematoxylin-eosin(HE)staining.The homocysteine(Hcy)in serum was detected by enzyme linked immunosorbent assay(ELISA).TdT-mediated dUTP nick-end labeling(TUNEL)was used to detect apoptosis in retinal tissue.Evans blue dye was used to detect the permeability of retinal blood vessels.The platelet endothelial cell adhesion molecule-1(CD31)and vascular endothelial growth factor receptor(VEGFR)protein were detected by Western blot.The 3-nitrotyrosine(3-NT)and 4-hydroxynonanine(4-HNE)were detected by immunohistochemistry.RESULTS:The density of retinal blood vessels,retinal thickness,retinal vascular permeability and the proportion of apoptotic cells of retinal tissue in the model group increased significantly than control group(P<0.05).The Hcy in serum and the levels of CD31,VEGFR,3-NT,and 4-HNE in retinal tissue increased significantly in the model group(P<0.01).Folic acid and decitabine both reversed these changes significantly,and the combination of the folic acid and decitabine worked best.CONCLUSION:The combination of folic acid and decitabine has a more significant protective effect on the retina in diabetic mice.

Seabuckthorn juice alleviates allergic symptoms in shrimp-induced food allergy mice

Tropomyosin(TM)in shrimp is one of the predominant causes of food allergy around the world.In the present study,the effect of seabuckthorn juice against TM-induced shrimp allergy was investigated in BALB/c mice.Allergic symptoms,spleen index,intestinal section and diarrhea were measured in shrimp allergy mice.As the results,seabuckthorn juice suppressed the lesions in jejunum tissue,diarrhea and allergic symptoms in shrimp allergy mice.Seabuckthorn juice also reduced serum concentrations of tumor necrosis factor-a(TNF-α)as well as immunoglobulin E(IgE)and stimulated the secretion of interleukin-10(IL-10)in mice with shrimp allergy.Taken together,our findings suggest that increased IL-10 by seabuckthorn juice inhibits Th2 cytokine production to suppress shrimp allergic symptoms.Furthermore,seabuckthorn juice also regulates shrimp allergy by reducing jejunum lesions,inhibiting levels of TNF-αand IgE.

Exploring the regulatory mechanism of tRNA-derived fragments 36 in acute pancreatitis based on small RNA sequencing and experiments

BACKGROUND Acute pancreatitis(AP)is a disease featuring acute inflammation of the pancreas and histological destruction of acinar cells.Approximately 20%of AP patients progress to moderately severe or severe pancreatitis,with a case fatality rate of up to 30%.However,a single indicator that can serve as the gold standard for prognostic prediction has not been discovered.Therefore,gaining deeper insights into the underlying mechanism of AP progression and the evolution of the disease and exploring effective biomarkers are important for early diagnosis,progression evaluation,and precise treatment of AP.AIM To determine the regulatory mechanisms of tRNA-derived fragments(tRFs)in AP based on small RNA sequencing and experiments.METHODS Small RNA sequencing and functional enrichment analyses were performed to identify key tRFs and the potential mechanisms in AP.Reverse transcription quantitative polymerase chain reaction(RT-qPCR)was conducted to determine tRF expression.AP cell and mouse models were created to investigate the role of tRF36 in AP progression.Lipase,amylase,and cytokine levels were assayed to examine AP progression.Ferritin expression,reactive oxygen species,malondialdehyde,and ferric ion levels were assayed to evaluate cellular ferroptosis.RNA pull down assays and methylated RNA immunoprecipitation were performed to explore the molecular mechanisms.RESULTS RT-qPCR results showed that tRF36 was significantly upregulated in the serum of AP patients,compared to healthy controls.Functional enrichment analysis indicated that target genes of tRF36 were involved in ferroptosisrelated pathways,including the Hippo signaling pathway and ion transport.Moreover,the occurrence of pancreatic cell ferroptosis was detected in AP cells and mouse models.The results of interference experiments and AP cell models suggested that tRF-36 could promote AP progression through the regulation of ferroptosis.Furthermore,ferroptosis gene microarray,database prediction,and immunoprecipitation suggested that tRF-36 accelerated the progression of AP by recruiting insulin-like growth factor 2 mRNA binding protein 3(IGF2BP3)to the p53 mRNA m6A modification site by binding to IGF2BP3,which enhanced p53 mRNA stability and promoted the ferroptosis of pancreatic follicle cells.CONCLUSION In conclusion,regulation of nuclear pre-mRNA domain-containing protein 1B promoted AP development by regulating the ferroptosis of pancreatic cells,thereby acting as a prospective therapeutic target for AP.In addition,this study provided a basis for understanding the regulatory mechanisms of tRFs in AP.