Hepatocyte growth factor enhances the ability of dental pulp stem cells to ameliorate atherosclerosis in apolipoprotein E-knockout mice

BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.

Expression changes of microtubule associated protein 1B in the brain of Fmr1 knockout mice

Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in the brains of 1-week and 6-week old fragile X mental retardation-1 （FmrI） knockout （KO） mice were investigated by immunohistochemistry, Western blot, and in situ hybridization, with the age-matched wild type mice （WT） as controls. Results The mean optical density （MOD） of MAP1B was significantly decreased in each brain region in KO6W compared with WT6W, whereas in KO1W, this decrease was only found in the hippocampus and cerebellum. MAP1B in 6-week mice was much less than that in 1-week mice of the same genotype. The results of Western blot and in situ hybridization showed that MAP1B protein and MAP1B mRNA were significantly decreased in the hippocampus of both KO1W and KO6W. Conclusion The decreased MAP1B protein and MAP1B mRNA in the Fmrl knockout mice indicate that FMRP may positively regulate the expression of MAP1B.

Helicobacter pylori arginase mutant colonizes arginase Ⅱ knockout mice

AIM: To investigate the role of host and bacterial arginases in the colonization of mice by Helicobacter pylori (H.pylori).METHODS: H.pylori produces a very powerful urease that hydrolyzes urea to carbon dioxide and ammonium,which neutralizes acid.Urease is absolutely essential to H.pylori pathogenesis;therefore,the urea substrate must be in ample supply for urease to work efficiently.The urea substrate is most likely provided by arginase activity,which hydrolyzes L-arginine to L-ornithine and urea.Previous work has demonstrated that H.pylori arginase is surprisingly not required for colonization of wild-type mice.Hence,another in vivo source of the critical urea substrate must exist.We hypothesized that the urea source was provided by host arginase Ⅱ,since this enzyme is expressed in the stomach,and H.pylori has previously been shown to induce the expression of murine gastric arginase Ⅱ.To test this hypothesis,wild-type and arginase (rocF) mutant H.pylori strain SS1 were inoculated into arginase Ⅱ knockout mice.RESULTS: Surprisingly,both the wild-type and rocF mutant bacteria still colonized arginase Ⅱ knockout mice.Moreover,feeding arginase Ⅱ knockout mice the host arginase inhibitor S-(2-boronoethyl)L-cysteine (BEC),while inhibiting > 50% of the host arginase Ⅰ?activity in several tissues,did not block the ability of the rocF mutant H.pylori to colonize.In contrast,BEC poorly inhibited H.pylori arginase activity.CONCLUSION: The in vivo source for the essential urea utilized by H.pylori urease is neither bacterial arginase nor host arginase Ⅱ;instead,either residual host arginase Ⅰ?or agmatinase is probably responsible.

Treadmill exercise in combination with acousto-optic and olfactory stimulation improves cognitive function in APP/PS1 mice through the brain-derived neurotrophic factor-and Cygb-associated signaling pathways

A reduction in adult neurogenesis is associated with behavioral abnormalities in patients with Alzheimer's disease.Consequently,enhancing adult neurogenesis represents a promising therapeutic approach for mitigating disease symptoms and progression.Nonetheless,nonpharmacological interventions aimed at inducing adult neurogenesis are currently limited.Although individual non-pharmacological interventions,such as aerobic exercise,acousto-optic stimulation,and olfactory stimulation,have shown limited capacity to improve neurogenesis and cognitive function in patients with Alzheimer's disease,the therapeutic effect of a strategy that combines these interventions has not been fully explored.In this study,we observed an age-dependent decrease in adult neurogenesis and a concurrent increase in amyloid-beta accumulation in the hippocampus of amyloid precursor protein/presenilin 1 mice aged 2-8 months.Amyloid deposition became evident at 4 months,while neurogenesis declined by 6 months,further deteriorating as the disease progressed.However,following a 4-week multifactor stimulation protocol,which encompassed treadmill running(46 min/d,10 m/min,6 days per week),40 Hz acousto-optic stimulation(1 hour/day,6 days/week),and olfactory stimulation(1 hour/day,6 days/week),we found a significant increase in the number of newborn cells(5'-bromo-2'-deoxyuridine-positive cells),immature neurons(doublecortin-positive cells),newborn immature neurons(5'-bromo-2'-deoxyuridine-positive/doublecortin-positive cells),and newborn astrocytes(5'-bromo-2'-deoxyuridine-positive/glial fibrillary acidic protein-positive cells).Additionally,the amyloid-beta load in the hippocampus decreased.These findings suggest that multifactor stimulation can enhance adult hippocampal neurogenesis and mitigate amyloid-beta neuropathology in amyloid precursor protein/presenilin 1 mice.Furthermore,cognitive abilities were improved,and depressive symptoms were alleviated in amyloid precursor protein/presenilin 1 mice following multifactor stimulation,as evidenced by Morris water maze,novel object recognition,forced swimming test,and tail suspension test results.Notably,the efficacy of multifactor stimulation in consolidating immature neurons persisted for at least 2weeks after treatment cessation.At the molecular level,multifactor stimulation upregulated the expression of neuron-related proteins(NeuN,doublecortin,postsynaptic density protein-95,and synaptophysin),anti-apoptosis-related proteins(Bcl-2 and PARP),and an autophagyassociated protein(LC3B),while decreasing the expression of apoptosis-related proteins(BAX and caspase-9),in the hippocampus of amyloid precursor protein/presenilin 1 mice.These observations might be attributable to both the brain-derived neurotrophic factor-mediated signaling pathway and antioxidant pathways.Furthermore,serum metabolomics analysis indicated that multifactor stimulation regulated differentially expressed metabolites associated with cell apoptosis,oxidative damage,and cognition.Collectively,these findings suggest that multifactor stimulation is a novel non-invasive approach for the prevention and treatment of Alzheimer's disease.

Type Ⅰ interferon receptor knockout mice as models for infection of highly pathogenic viruses with outbreak potential

Due to their inability to generate a complete immune response, mice knockout for type I interferon （IFN） receptors （Ifnar-/-） are more susceptible to viral infections, and are thus commonly used for pathogenesis studies. This mouse model has been used to study many diseases caused by highly pathogenic viruses from many families, including the Flaviviridae, Filoviridae, Arenaviridae, Bunyaviridae, Henipaviridae, and Togaviridae. In this review, we summarize the findings from these animal studies, and discuss the pros and cons of using this model versus other known methods for studying pathogenesis in animals.

Effects of P301L-TAU on post-translational modifications of microtubules in human iPSC-derived cortical neurons and TAU transgenic mice

TAU is a microtubule-associated protein that promotes microtubule assembly and stability in the axon.TAU is missorted and aggregated in an array of diseases known as tauopathies.Microtubules are essential for neuronal function and regulated via a complex set of post-translational modifications,changes of which affect microtubule stability and dynamics,microtubule interaction with other proteins and cellular structures,and mediate recruitment of microtubule-severing enzymes.As impairment of microtubule dynamics causes neuronal dysfunction,we hypothesize cognitive impairment in human disease to be impacted by impairment of microtubule dynamics.We therefore aimed to study the effects of a disease-causing mutation of TAU(P301L)on the levels and localization of microtubule post-translational modifications indicative of microtubule stability and dynamics,to assess whether P301L-TAU causes stability-changing modifications to microtubules.To investigate TAU localization,phosphorylation,and effects on tubulin post-translational modifications,we expressed wild-type or P301L-TAU in human MAPT-KO induced pluripotent stem cell-derived neurons(i Neurons)and studied TAU in neurons in the hippocampus of mice transgenic for human P301L-TAU(p R5 mice).Human neurons expressing the longest TAU isoform(2N4R)with the P301L mutation showed increased TAU phosphorylation at the AT8,but not the p-Ser-262 epitope,and increased polyglutamylation and acetylation of microtubules compared with endogenous TAU-expressing neurons.P301L-TAU showed pronounced somatodendritic presence,but also successful axonal enrichment and a similar axodendritic distribution comparable to exogenously expressed 2N4R-wildtype-TAU.P301L-TAU-expressing hippocampal neurons in transgenic mice showed prominent missorting and tauopathy-typical AT8-phosphorylation of TAU and increased polyglutamylation,but reduced acetylation,of microtubules compared with non-transgenic littermates.In sum,P301L-TAU results in changes in microtubule PTMs,suggestive of impairment of microtubule stability.This is accompanied by missorting and aggregation of TAU in mice but not in i Neurons.Microtubule PTMs/impairment may be of key importance in tauopathies.

Investigating Müller glia reprogramming in mice: a retrospective of the last decade, and a look to the future

Müller glia,as prominent glial cells within the retina,plays a significant role in maintaining retinal homeostasis in both healthy and diseased states.In lower vertebrates like zebrafish,these cells assume responsibility for spontaneous retinal regeneration,wherein endogenous Müller glia undergo proliferation,transform into Müller glia-derived progenitor cells,and subsequently regenerate the entire retina with restored functionality.Conversely,Müller glia in the mouse and human retina exhibit limited neural reprogramming.Müller glia reprogramming is thus a promising strategy for treating neurodegenerative ocular disorders.Müller glia reprogramming in mice has been accomplished with remarkable success,through various technologies.Advancements in molecular,genetic,epigenetic,morphological,and physiological evaluations have made it easier to document and investigate the Müller glia programming process in mice.Nevertheless,there remain issues that hinder improving reprogramming efficiency and maturity.Thus,understanding the reprogramming mechanism is crucial toward exploring factors that will improve Müller glia reprogramming efficiency,and for developing novel Müller glia reprogramming strategies.This review describes recent progress in relatively successful Müller glia reprogramming strategies.It also provides a basis for developing new Müller glia reprogramming strategies in mice,including epigenetic remodeling,metabolic modulation,immune regulation,chemical small-molecules regulation,extracellular matrix remodeling,and cell-cell fusion,to achieve Müller glia reprogramming in mice.

Truncating PICK1 Variant Identified in Azoospermia Affected Mitochondrial Dysfunction in Knockout Mice

Objective The protein interacting with C kinase 1(PICK1)plays a critical role in vesicle trafficking,and its deficiency in sperm cells results in abnormal vesicle trafficking from Golgi to acrosome,which eventually disrupts acrosome formation and leads to male infertility.Methods An azoospermia sample was filtered,and the laboratory detection and clinical phenotype indicated typical azoospermia in the patient.We sequenced all of the exons in the PICK1 gene and found that there was a novel homozygous variant in the PICK1 gene,c.364delA(p.Lys122SerfsX8),and this protein structure truncating variant seriously affected the biological function.Then we constructed a PICK1 knockout mouse model using clustered regularly interspaced short palindromic repeat cutting technology(CRISPRc).Results The sperm from PICK1 knockout mice showed acrosome and nucleus abnormalities,as well as dysfunctional mitochondrial sheath formation.Both the total sperm and motility sperm counts were decreased in the PICK1 knockout mice compared to wild-type mice.Moreover,the mitochondrial dysfunction was verified in the mice.These defects in the male PICK1 knockout mice may have eventually led to complete infertility.Conclusion The c.364delA novel variant in the PICK1 gene associated with clinical infertility,and pathogenic variants in the PICK1 may cause azoospermia or asthenospermia by impairing mitochondrial function in both mice and humans.

AAV mediated carboxyl terminus of Hsp70 interacting protein overexpression mitigates the cognitive and pathological phenotypes of APP/PS1 mice

The E3 ubiquitin ligase,carboxyl terminus of heat shock protein 70(Hsp70)interacting protein(CHIP),also functions as a co-chaperone and plays a crucial role in the protein quality control system.In this study,we aimed to investigate the neuroprotective effect of overexpressed CHIP on Alzheimer’s disease.We used an adeno-associated virus vector that can cross the blood-brain barrier to mediate CHIP overexpression in APP/PS1 mouse brain.CHIP overexpression significantly ameliorated the performance of APP/PS1 mice in the Morris water maze and nest building tests,reduced amyloid-βplaques,and decreased the expression of both amyloid-βand phosphorylated tau.CHIP also alleviated the concentration of microglia and astrocytes around plaques.In APP/PS1 mice of a younger age,CHIP overexpression promoted an increase in ADAM10 expression and inhibitedβ-site APP cleaving enzyme 1,insulin degrading enzyme,and neprilysin expression.Levels of HSP70 and HSP40,which have functional relevance to CHIP,were also increased.Single nuclei transcriptome sequencing in the hippocampus of CHIP overexpressed mice showed that the lysosomal pathway and oligodendrocyte-related biological processes were up-regulated,which may also reflect a potential mechanism for the neuroprotective effect of CHIP.Our research shows that CHIP effectively reduces the behavior and pathological manifestations of APP/PS1 mice.Indeed,overexpression of CHIP could be a beneficial approach for the treatment of Alzheimer’s disease.

Gamma-glutamyl transferase 5 overexpression in cerebrovascular endothelial cells improves brain pathology,cognition,and behavior in APP/PS1 mice

In patients with Alzheimer’s disease,gamma-glutamyl transferase 5(GGT5)expression has been observed to be downregulated in cerebrovascular endothelial cells.However,the functional role of GGT5 in the development of Alzheimer’s disease remains unclear.This study aimed to explore the effect of GGT5 on cognitive function and brain pathology in an APP/PS1 mouse model of Alzheimer’s disease,as well as the underlying mechanism.We observed a significant reduction in GGT5 expression in two in vitro models of Alzheimer’s disease(Aβ_(1-42)-treated hCMEC/D3 and bEnd.3 cells),as well as in the APP/PS1 mouse model.Additionally,injection of APP/PS1 mice with an adeno-associated virus encoding GGT5 enhanced hippocampal synaptic plasticity and mitigated cognitive deficits.Interestingly,increasing GGT5 expression in cerebrovascular endothelial cells reduced levels of both soluble and insoluble amyloid-βin the brains of APP/PS1 mice.This effect may be attributable to inhibition of the expression ofβ-site APP cleaving enzyme 1,which is mediated by nuclear factor-kappa B.Our findings demonstrate that GGT5 expression in cerebrovascular endothelial cells is inversely associated with Alzheimer’s disease pathogenesis,and that GGT5 upregulation mitigates cognitive deficits in APP/PS1 mice.These findings suggest that GGT5 expression in cerebrovascular endothelial cells is a potential therapeutic target and biomarker for Alzheimer’s disease.

Effects of SIRT1 Gene Knock-Out via Activation of SREBP2 Protein-Mediated PI3K/AKT Signaling on Osteoarthritis in Mice

This study investigated the effects of SIRT1 gene knock-out on osteoarthritis in mice, and the possible roles of SREBP2 protein and the PI3K/AKT signaling pathway in the effects. Mice were randomly divided into a normal group and a SIRT1 gene knock-out group（6 mice in each group）. In these groups, one side of the knee anterior cruciate ligament was traversed, and the ipsilateral medial meniscus was cut to establish an osteoarthritis model of knee joint. The countralateral synovial bursa was cut out, serving as controls. The knee joint specimens were then divided into four groups： SIRT1^（＋/＋） control group（group A, n=6）; SIRT1^（＋/＋） osteoarthritis group（group B, n=6）; SIRT1^（–/–） control group（group C, n=6）; SIRT1^（–/–） osteoarthritis group（group D, n=6）. HE staining, Masson staining, Safranin O-Fast Green staining and Van Gieson staining were used to observe the morphological changes in the articular cartilage of the knee. Immunohistochemical staining was employed to detect the expression of SIRT1, SREBP2, VEGF, AKT, HMGCR and type Ⅱ collagen proteins. SA-β-gal staining was utilized to evaluate chondrocyte aging. The results showed clear knee joint cartilage destruction and degeneration in the SIRT1^（–/–） osteoarthritis group. The tidal line was twisted and displaced anteriorly. Type Ⅱ collagen was destroyed and distributed unevenly. Compared with the SIRT1^（＋/＋） osteoarthritis group and SIRT1^（–/–） control group, SIRT1 protein expression was not obviously changed in the SIRT1^（–/–） osteoarthritis group（P〉0.05）, while the expression levels of the SREBP2, VEGF and HMGCR proteins were significantly increased（P〈0.05） and the levels of AKT and type Ⅱ collagen proteins were significantly decreased（P〈0.05）. SIRT1 gene knock-out may aggravate cartilage degeneration in osteoarthritis by activating the SREBP2 protein-mediated PI3K/AKT signalling pathway, suggesting that SIRT1 gene may play a protective role against osteoarthritis.

Molecular mechanism about lymphogenous metastasis of hepatocarcinoma cells in mice

AIM: To investigate the correlation between lymphogenous metastasis and matrix metalloproteinases (MMPs) activity and the expression of Fas ligand of tumor cells in lymph nodes. METHODS: Fifty-six inbred 615-mice were equally divided into 2 groups and inoculated with Hca-F and Hca-P cells. Their lymph node metastatic rates were examined. Growth fraction of lymphocytes in host lymph nodes was detected by flow cytometry. The Hca-F and Hca-P cells were cultured with extract of lymph node, liver or spleen. The quantity of MMPs in these supernatants was examined by zymographic analysis. The expression of Fas ligand, PCNA, Bcl-2 protein of Hca-F and Hca-P cells in the mice were examined by immunohistochemistry. The apoptosis signals of macro-phages in lymph nodes were observed with in situ DNA fragmentation. RESULTS: On the 28th day post-inoculation, the lymph node metastatic rate of HcaF was 80%(16/20), whereas that of Hca-P was 25%(5/20). The growth fraction of lymphocytes was as follows: in the Hca-F cells, the proliferating peak of lymphocytes appeared on the 14th day post inoculation and then decreased rapidly, while in HcaP cells, the peak appeared on the 7th day post inoculation and then kept at a high level. With the extract of lymph node, the quantity of the MMP-9 activity increased (P【0.01) and active MMP-9 and MMP-2 were produced by both Hca-F and Hca-P tumor cells, which did not produce MMPs without the extract of lymph node or with the extracts of the liver and spleen. The expression of Fas Ligand of Hca-F cells was stronger than that of Hca-P cells (P 【0.01). The expressions of PCNA and Bcl-2 protein of Hca-F cells in the tumors of inoculated area were the same as that of Hca-P cells. In situ DNA fragmentation showed that the positive signals of macrophages were around Hca-F cells. CONCLUSION: Secretion of MMPs which was associated with metastatic ability of Hca-F and Hca-P tumor cells depends on the environment of lymph nodes. The increased expression of Fas ligand protein of Hca-F tumor cells with high lymphogenous metastatic potential in lymph nodes may help tumor cells escape from being killed by host lymphocytes.

Tanshinone Ⅱ-A Attenuates and Stabilizes Atherosclerotic Plaques in Apolipoprotein E Knockout Mice Fed with High Cholesterol Diet

Objective TanshinoneⅡ-A(Tan),a bioactive diterpene isolated fromSalvia miltiorrhiza Bunge(Danshen),possesses anti-oxidant and anti-in-flammatory activities.The present study investigated whether Tan can reduce and stabilize atherosclerotic plaques in Apolipoprotein E knockout(ApoE-/-) mice maintained on a high cholesterol diet(HCD).Methods and Results Six week-old mice challenged with HCD were ran-domly assigned to 4 groups: C57BL/6J,ApoE-/-,ApoE-/-+30 mg/kg.d Tan and ApoE-/-+10 mg/kg.d Tan.After 16 weeks of inter-vention,Tan treated mice showed decreased atherosclerotic lesion size in the aortic sinus and face aorta.Furthermore,immunohistochemical a-nalysis revealed that Tan rendered the lesion composition a more stable phenotype as evidenced by reduced necrotic cores,decreased macrophageinfiltration,increased smooth muscle cell and collagen content.Tan also significantly reduced in situ superoxide anion production,aortic expres-sion of NF-κB,and matrix metalloproteinase-9(MMP-9).In vitro treatment of RAW264.7 macrophages with Tan significantly suppressed oxi-dized LDL-induced reactive oxygen species production,pro-inflammatory cytokine(IL-6,TNF-α,MCP-1) expression,and MMP-9 activity.Conclusions Tan attenuates the development of atherosclerotic lesions and promotes plaque stability in ApoE-/-mice by reducing vascular oxi-dative stress and inflammatory responses.Our findings highlightTan as a potential therapeutic agentto preventatherosclerotic cardiovascular dis-eases.

1H-NMR-based Metabolomic approach to evaluating total flavonoids of Ocimum Basilicum Linn in apolipomtein E knockout mice

OBJECTIVE To observe the effects of serum metabolites by using ~1H-NMR-based metabonomic approach to explore the possible mechanisms of total flavonoids in Ocimum BasilicumLinn(OBL) on atherosclerosis in apolipomtein E knockout(ApoE-/-) mice.METHODS Six-week-old male apoE knockout mice were divided into four groups(n=10) and fed with high fet diet:model,Simv.astatin,OBL-H,OBL-M and OBL-L groups.The homogeneous male mice of C57 BL/6 J were used as the normol group and fed with normal chow diet.After 14 weeks,~1H-NMR technology was used to ex.plore the variability of serum metabolites by the method of PLS-DA and OPLS-DA.RESULTS Com.pared with normal group,Model group showed a significant increase in the serum levels of VLDL,LDL,β-hydroxyisobutyrate,lactate,myo-inositol and showed a significant decrease in the serum levels of al.anine,glutamine,proline,carnitine,methylamine,citrate,creatine,choline,taurine,pyruvate,β-glu.cose,α-glucose,glycine,lysine.Combined with model group OBL-H,OBL-M,OBL-L groups showed the effects of regulating the levels of different metabolites of the glucose,lipid and amino acid metabo.lism.CONCLUSION The anti-atheros-clerotic activity of total flavonoids in Ocimum BasilicumLinn may be related not only to regulation of lipid metabolism,but also glycometabolism and amino acid metabolism.

Triptolide prolonged allogeneic islet graft survival in chemically induced and spontaneously diabetic mice without impairment of islet function

BACKGROUND: Triptolide (TPT) is a diterpenoid triepoxide extracted from the Chinese herb Tripterygium wilfordii Hook. F. It exhibits potent immunosuppressive and anti-inflammatory properties. This study was undertaken to investigate its effects on prolongation of islet allograft survival in rodents. Additionally, we investigated whether TPT would be toxic to islet function in vivo. METHODS: We transplanted BALB/c islets to either chemically induced diabetic C57BL/6 mice or spontaneously diabetic non-obese diabetic (NOD) mice. TPT was injected within 2 weeks or continuously, until rejection, in the two combinations. Then, we evaluated the toxicity of TPT on islet function by daily injection to naive BALB/c or diabetic BALB/c that was cured by syngeneic islet transplantation under the kidney capsule. Mice injected with cyclosporine A (CsA) or vehicle served as controls. Intraperitoneal glucose tolerance tests (IPGTTs) performed at 4 and 8 weeks in the naive BALB/c group, and at 2, 4, 6, and 8 weeks in the syngeneic transplanted group. RESULTS: The medium survival time of islets allograft from TPT treated C57BL/6 and NOD recipients were 28.5 days (range 24-30 days, n=10) and 33.0 days (range 15-47 days, n=6), respectively, and they were significantly different from those of the vehicle treated controls, which were 14.0 days (range 13-16 days, n=6) and 5.0 days (range 4-10 days, n=6), respectively (all P<0.0001). The IPGTT demonstrated that there was no difference between the TPT treated and vehicle treated groups, either in the normal or syngeneic transplanted islet BALB/c mice. However, CsA injection impaired islet function in both normal and syngeneic transplanted mice as early as 4 weeks. CONCLUSION: TPT prolonged islets allograft survival in a chemically induced diabetic or an autoimmune diabetic murine model without impairment of islet function. (Hepatobiliary Pancreat Dis Int 2010; 9: 312-318)

Effects of estradiol-17β and bisphenol A administered chronically to mice throughout pregnancy and lactation on the male pups＇ reproductive system

Aim： To assess the effect of estradiol-17β （E2） and bisphenol A （BPA） administered chronically by implanting a silicone tube throughout pregnancy and lactation on male pups＇ reproductive system in ICR mice. Methods： Female mice were implanted with a tube filled with 10 ng, 500 ng, 1 μg, or 10 μg of E2, or 100 μg or 5 mg of BPA, before mating. The tube was kept in the mice throughout pregnancy and lactation, until the pups had weaned at 4 weeks of age. During the period, E2 was released from the tube at 120 pg or 6, 12 or 120 ng/day, and BPA at 1.2 or 60 μg/day. Results： Most of the mice given 1 μg and 10 lag of E2 did not maintain their pregnancy. However, the other groups showed high rates of birth, more than 70%. At age of 4 weeks, the male pups were killed. Body weight and reproductive organ weights （testes, epididymides and accessory reproductive glands） in the treated groups did not differ from the control values, whereas the percentage of seminiferous tubules in the testis with mature spermatids was significantly lower in the groups given 10 ng and 500 ng of E2 and 5 mg of BPA than that in the control. Conclusion： Chronic exposure to E2 and BPA might disrupt spermatogenesis in male pups. （Asian JAndrol 2008 Mar; 10： 271-276）

Testis-specific Fankl gene in knockdown mice produces oligospermia via apoptosis

Fankl is exclusively expressed in the testis from the meiosis phase to the haploid phase of spermatogenesis. In this study, we examined the function of Fankl by establishing a Fankl-knockdown transgenic mouse model. The apoptotic statuses of the testes of the transgenic mice were tested using the terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） method. The FANK1 consensus DNA-binding sequence was identified using cyclic amplification of sequence target （CAST） analysis. Differentially expressed genes were examined using microarray analysis. A reduction in sperm number and an increase in apoptotic spermatocytes were observed in Fankl-knockdown mice, and the apoptotic cells were found to be primarily spermatogonia and spermatocytes. The CAST results demonstrated that the consensus DNA-binding sequence was AAAAAG, in which the percentage occurrence of each base at each position ranged from 55 to 86%. This sequence was present in the promoter regions of 10 differentially expressed genes that were examined using microarray analysis. In total, 17 genes were differentially expressed with changes in their expression levels greater than twofold. The abnormal expression of Fankl target genes that were regulated directly or indirectly by Fankl reduced the number of sperm in the knockdown mice. Thus, FANK1 may Dlav a pivotal role in sDermato^enesis as a transcription factor.Fankl is exclusively expressed in the testis from the meiosis phase to the haploid phase of spermatogenesis. In this study, we examined the function of Fankl by establishing a Fankl-knockdown transgenic mouse model. The apoptotic statuses of the testes of the transgenic mice were tested using the terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） method. The FANK1 consensus DNA-binding sequence was identified using cyclic amplification of sequence target （CAST） analysis. Differentially expressed genes were examined using microarray analysis, A reduction in sperm number and an increase in apoptotic spermatocytes were observed in Fankl-knockdown mice, and the apoptotic cells were found to be primarily spermatogonia and spermatocytes. The CAST results demonstrated that the consensus DNA-binding sequence was AAAAAG, in which the percentage occurrence of each base at each position ranged from 55 to 86%. This sequence was present in the promoter regions of 10 differentially expressed genes that were examined using microarray analysis. In total, 17 genes were differentially expressed with changes in their expression levels greater than twofold. The abnormal expression of Fankl target genes that were regulated directly or indirectly by Fankl reduced the number of sperm in the knockdown mice. Thus, FANK1 may play a pivotal role in spermatogenesis as a transcription factor.

Di-(n-butyl)-phthalate-induced Oxidative Stress and Depression-like Behavior in Mice with or without Ovalbumin Immunization

Objective To investigate the relationship between atopic allergy and depression and the role of DBP in the development of depression. Methods BALB/c mice were randomly divided into eight groups：saline;ovalbumin （OVA）-immunized;saline＋DBP （0.45 mg/kg·183;d）; saline＋DBP （45 mg/kg·d）; DBP （0.45 mg/kg·d） OVA-immunized; DBP （45 mg/kg·d） OVA-immunized; saline＋hydrocortisone （30 mg/kg·d）; and hydrocortisone （30 mg/kg·d）-exposed OVA-immunized. Behavior （e.g. open-field, tail suspension, and forced swimming tests）, viscera coefficients （brain and spleen）, oxidative damage [e.g. reactive oxygen species （ROS）, malondialdehyde （MDA）, and glutathione （GSH）], as well as levels of IgE and IL-4, were then analyzed. Results In the saline and OVA groups, the degree of depression symptoms in mice increased with increasing DBP concentration. Additionally, the OVA-immunity groups were associated with more serious depressive behavior compared with the same exposure concentration in the saline group. Oxidative damage was associated with a dose-dependent increase in DBP in the different groups. IL-4 and IgE levels were associated with low-dose DBP stimulation, which changed to high-dose inhibition with increasing DBP exposure, possibly due to spleen injury seen at high DBP concentrations.Conclusion Development of an atopic allergy has the potential to increase the risk of depression in mice, and it seems that DBP helps OVA to exert its effect in our present model. Moreover, the results of our study implicate a certain connection between brain oxidative stress and depression, which deserves a further exploration.

Can outbred mice be used as a mouse model of mild cognitive impairment?

Deficits in spatial learning and memory are some of the earliest symptoms in mild cognitive impairment （MCI）. However, there are few valid MCI animal models available to evaluate putative therapeutic strategies. The aim of this study was to obtain a natural animal model of MCI. Outbred Kunming （aged 5 and 12.5 months） and ICR （7 and 12 months） mice were utilized in the present study. Morris water maze and radial six-arm water maze （RAWM） were simultaneously used to evaluate impaired spatial learning and memory in middle-aged mice （approximately 12 months of age）. Compared with younger mice in the respective groups, the middle-aged mice suffered visible impairment of spatial memory in the Morris water maze and RAWM, and mild spatial learning deficiency occurred in the RAWM study alone. Thus outbred Kunming and ICR mice could be utilized as a natural animal model for MCI, in particular for memory impairment studies.