ICR Strain Research Articles

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increased ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes.

Effects of growth factors during in vitro culture of mouse and rat embryos

The development of embryos of the ICR, HT1AN/lcgn, HT1AC/lcgn, and C57BL/6J-Ay mouse strains, as well as the OXYS/lcgn rat strains, in the culture in vitro was studied under the influence of the granulocyte- macrophage colony stimulating factor (GM-CSF) and epidermal growth factor (EGF). Both mice and rats embryos were initially frozen according to the standard protocol of the freezing program using glycerol and sucrose as cryoprotectant and culture in the rat 1-cell embryo culture medium (R1ECM) for either 24 h (mice) or 72 h (rats) after thawing. The effects of the growth factors were studied on 8-cell embryos in mice and 2–4-cell embryos in rats. The GM-CSF effect resulted in an increase in the percentage of developing embryos in the mice of both strains (HT1AC/lcgn and C57BL/6J-Ay); at the same time, no EGF effect on the mice embryos was found. The situation was the opposite for rats. The EGF effect resulted in the acceleration of the development until the stage of blastocyst in the OXYS/lcgn strain rats, but no GM-CSF effect on the rat embryos was found. The acceleration of the development is observed during the joint culture of 4-cell embryos of the HT1AN/lcgn mice strain with later stages of development (embryos at the morula stage) of the ICR strain. The results of the presented experiments indicate a species specificity of the effect of the growth factors on the mice and rats embryos, as well as demonstrating that joint culture of embryos of the later stages (morula) of mice with earlier stages (4-cell embryos) has a stimulating effect on the latter.

PKA-CREB-BDNF signaling regulated long lasting antidepressant activities of Yueju but not ketamine

Yueju confers antidepressant effects in a rapid and long-lasting manner, similar to ketamine. CREB (cAMP-response element binding protein) signaling is implicated in depression pathology and antidepressant responses. However, the role of CREB and associated brain derived neurotrophic factor (BDNF) signaling in rapid and long-lasting antidepressant effects remains unclear. Here, we demonstrated that ICR and Kunming strain mice conferred antidepressant responses lasting for 1 and 5 days, respectively, following a single dose of Yueju. One day post Yueju in Kunming but not ICR strain mice, expression of total and phosphorylated CREB, as well as the CREB signaling activator, PKA (protein kinase A) was up-regulated in the hippocampus. Although BDNF gene expression increased at 3 hours in both strains, it remained up-regulated at 1 day only in Kunming mice. Ketamine showed similar strain-dependent behavioral effects. However, blockade of PKA/CREB signaling blunted the antidepressant effects and reversed the up-regulation of BDNF gene expression by Yueju, but not ketamine. Conversely, blockade of mammalian target of rapamycin signaling led to opposite effects. Taken altogether, prolonged transcriptional up-regulation of hippocampal BDNF may account for the stain-dependent enduring antidepressant responses to Yueju and ketamine, but it was mediated via PKA/CREB pathway only for Yueju.

In vitro and in vivo studies of a potent capsid-binding inhibitor of enterovirus 71.

Enterovirus 71 (EV-A71) is an important pathogen that can cause severe neurological symptoms and even death. Our aim was to identify potent anti-EV-A71 compounds and study their underlying mechanisms and in vivo activity. We identified a potent imidazolidinone derivative (abbreviated to PR66) as an inhibitor of EV-A71 infection from the screening of compounds and subsequent structure-based modification. Time-course treatments and resistant virus selection of PR66 were employed to study the mode of mechanism of PR66. In vivo activity of PR66 was tested in the ICR strain of new-born mice challenged with EV-A71/4643/MP4. PR66 could impede the uncoating process during viral infection via interaction with capsid protein VP1, as shown by a resistant virus selection assay. Using site-directed mutagenesis, we confirmed that a change from valine to phenylalanine in the 179th amino acid residue of the cDNA-derived resistant virus resulted in resistance to PR66. PR66 increased the virion stability of WT viruses, but not the PR66-resistant mutant, in a particle stability thermal release assay. We further showed that PR66 had excellent anti-EV-A71 activity in an in vivo mouse model of disease, with a dose-dependent increase in survival rate and in protection against virus-induced hind-limb paralysis following oral or intraperitoneal administration. This was associated with reductions of viral titres in brain and muscle tissues. We demonstrated here for the first time that an imidazolidinone derivative (PR66) could protect against EV-A71-induced neurological symptoms in vivo by suppressing EV-A71 replication. This involved binding to and restricting viral uncoating.

MR Imaging–derived Oxygen-Hemoglobin Dissociation Curves and Fetal-Placental Oxygen-Hemoglobin Affinities

Purpose To generate magnetic resonance (MR) imaging-derived, oxygen-hemoglobin dissociation curves and to map fetal-placental oxygen-hemoglobin affinity in pregnant mice noninvasively by combining blood oxygen level-dependent (BOLD) T2* and oxygen-weighted T1 contrast mechanisms under different respiration challenges. Materials and Methods All procedures were approved by the Weizmann Institutional Animal Care and Use Committee. Pregnant mice were analyzed with MR imaging at 9.4 T on embryonic days 14.5 (eight dams and 58 fetuses; imprinting control region ICR strain) and 17.5 (21 dams and 158 fetuses) under respiration challenges ranging from hyperoxia to hypoxia (10 levels of oxygenation, 100%-10%; total imaging time, 100 minutes). A shorter protocol with normoxia to hyperoxia was also performed (five levels of oxygenation, 20%-100%; total imaging time, 60 minutes). Fast spin-echo anatomic images were obtained, followed by sequential acquisition of three-dimensional gradient-echo T2*- and T1-weighted images. Automated registration was applied to align regions of interest of the entire placenta, fetal liver, and maternal liver. Results were compared by using a two-tailed unpaired Student t test. R1 and R2* values were derived for each tissue. MR imaging-based oxygen-hemoglobin dissociation curves were constructed by nonlinear least square fitting of 1 minus the change in R2*divided by R2*at baseline as a function of R1 to a sigmoid-shaped curve. The apparent P50 (oxygen tension at which hemoglobin is 50% saturated) value was derived from the curves, calculated as the R1 scaled value (x) at which the change in R2* divided by R2*at baseline scaled (y) equals 0.5. Results The apparent P50 values were significantly lower in fetal liver than in maternal liver for both gestation stages (day 14.5: 21% ± 5 [P = .04] and day 17.5: 41% ± 7 [P < .0001]). The placenta showed a reduction of 18% ± 4 in mean apparent P50 values from day 14.5 to day 17.5 (P = .003). Reproduction of the MR imaging-based oxygen-hemoglobin dissociation curves with a shorter protocol that excluded the hypoxic periods was demonstrated. Conclusion MR imaging-based oxygen-hemoglobin dissociation curves and oxygen-hemoglobin affinity information were derived for pregnant mice by using 9.4-T MR imaging, which suggests a potential to overcome the need for direct sampling of fetal or maternal blood. Online supplemental material is available for this article.

Investigation of In Vivo Antipyretic Activity of Thottea dependens leaves

Introduction: Leaves of Thottea dependens have been used as folk medicine in Malaysia for the treatment of several conditions involving pain and inflammation with accompanying pyrexia. However, there is no scientific evidence for its effectiveness to treat fever. Hence, the purpose of this study was to evaluate the anti-pyretic activity of methanol (MeOH) and aqueous (Aq) extracts of T. dependens leaves in albino mice (ICR strain). Methods: Thirty-six albino mice of both sexes, randomized into six groups (n=6) were used. Group I served as the control and received only the vehicle, normal saline. Group II received paracetamol 150 mg/kg body weight (b.w.) orally, groups III and IV received 100 and 200 mg/kg b.w. of the MeOH extract while groups V and VI received 100 and 200 mg/kg b.w. of the Aq extract orally. Pyrexia was induced by a subcutaneous injection of 10 mL/kg b.w. of 15% (w/v) yeast suspended in 0.5% (w/v) methylcellulose solution. LD50 of both extracts were determined according to the method of Organisation for Economic Co-operation and Development guidelines. Phytochemical screening was carried out qualitatively for the presence of different class of biologically active compounds. Results: LD50 values for both extracts were found to be greater than 1000 mg/kg b.w. indicating the wide margin of safety of T. dependens leaves. The MeOH extract at 200 mg/kg b.w. showed statistically significant (P&lt;0.01) dose dependent reduction of brewer’s yeast induced pyrexia in albino mice. Flavonoids, saponins and phenolic compounds were found to be major biologically active phytoconstituents in MeOH extract. Conclusion: The study showed that T. dependens leaves possess significant anti-pyretic activity, thus justifying its ethnomedicinal use in Malaysia to treat fever.

In vivo antimalarial activity of the endophytic actinobacteria, Streptomyces SUK 10.

Endophytic bacteria, such as Streptomyces, have the potential to act as a source for novel bioactive molecules with medicinal properties. The present study was aimed at assessing the antimalarial activity of crude extract isolated from various strains of actinobacteria living endophytically in some Malaysian medicinal plants. Using the four day suppression test method on male ICR strain mice, compounds produced from three strains of Streptomyces (SUK8, SUK10, and SUK27) were tested in vivo against Plasmodium berghei PZZ1/100 in an antimalarial screen using crude extracts at four different concentrations. One of these extracts, isolated from Streptomyces SUK10 obtained from the bark of Shorea ovalis tree, showed inhibition of the test organism and was further tested against P. berghei-infected mice for antimalarial activity at different concentrations. There was a positive relationship between the survival of the infected mouse group treated with 50 µg/kg body weight (bw) of ethyl acetate-SUK10 crude extract and the ability to inhibit the parasites growth. The parasite inhibition percentage for this group showed that 50% of the mice survived for more than 90 days after infection with the parasite. The nucleotide sequence and phylogenetic tree suggested that Streptomyces SUK10 may constitute a new species within the Streptomyces genus. As part of the drug discovery process, these promising finding may contribute to the medicinal and pharmaceutical field for malarial treatment.

Effects of growth factors during in vitro culture of mouse and rat embryos

In vitro culture of preimplantation embryos of ICR, HT1AN/Icgn, HT1AC/Icgn and C57BL/6J-Ay mouse strains as well as in OXYS/Icgn rat strain in media containing granulocyte-macrophage colony stimulating factor (GM-CSF) or epidermal growth factor (EGF) has been studied. Both mouse and rat embryos were first frozen in a programmable freezer after a standard protocol using a mixture of glycerol and sucrose as cryoprotectants, thawed and cultured in vitro in R1ECM (rat one-cell embryo culture medium) for 24 hours (mice) and 72 hours (rats). For the in vitro culture experiments with these growth factors, 8-cell frozen-thawed mouse embryos and 2–4-cell frozen-thawed rat embryos were used. Supplementation of the culture medium with GM-CSF improved the rate of embryonic development in HT1AC/Icgn and C57BL/6J-Ay strain mice, while EGF had no effect. The reverse was true of the rats. Supplementation of the culture medium with EGF increased the percentage of deve­loping blastocysts in OXYS/Icgn rat strain, while GM-CSF had no effect. Co-culture of four-cell embryos of HT1AN/ Icgn strain mice with more advanced embryonic stages (morulas) of a different strain ICR led to the facilitation preimplantation embryo development. Experimental results presented here reveal the species-specific effects of growth factors on mouse and rat embryos and indicate that co-culture of different stages of embryo development have stimulatory effects on earlier stages.

Oral treatment with herbal formula B401 alleviates penile toxicity in aging mice with manganism.

The present study aims to elucidate the roles of nitric oxide synthase activity, oxidative stress, inflammation, and apoptosis in penile toxicity of aging mice associated with excess manganese (Mn) treatment and to investigate the effect of oral treatment with the herbal formula B401 in this respect. ICR strain mice were divided into two groups: the vehicle (sham group) and the B401 (50 mg/kg) group. The mice were orally treated for 5 days; then a high single dose of MnCl2 (100 mg/kg) was given by intraperitoneal injection to the mice. One day after MnCl2 treatment, corpora cavernosal tissues of both Mn-treated mice and their controls were simultaneously sampled to examine their immunohistochemical staining and Western blot analysis. Nitric oxide (NO) production, levels of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS), expression levels of factors governing angiogenesis (vascular endothelial growth factor), oxidative stress (catalase, superoxide dismutase 2,4-hydroxynonenal), inflammation (tumor necrosis factor alpha), apoptosis (B-cell lymphoma 2 [Bcl-2], Bcl-2-associated X protein [Bax], cleaved poly(adenosine diphosphate-ribose) polymerase [c-PARP], cytochrome C, caspase-12, and caspase-3) were evaluated in penile corpus cavernosum of the mice. We found that penile toxicity in the mice was enhanced under excess Mn treatment through reduction of NOS activity and increase in oxidative stress, inflammation, and apoptosis in the penile cavernous tissue. Furthermore, the penile toxicity in mice with manganism was alleviated by oral B401 treatment through enhancement of both nitric oxide synthesis and angiogenesis, with simultaneous reduction of oxidative stress, inflammation, and apoptosis in penile corpus cavernosum. We suggest that the herbal formula B401 may serve as a potential dietotherapeutic supplement for penile toxicity or dysfunction in aging males.

New inhibitors of glucosylceramide synthase and their effect on cell fate

Abstract Glucosylceramide (GlcCer) is an essential glycosylated lipid found in organisms ranging from fungi to mammals. It is composed of a hydrophilic β-linked glucose and a hydrophobic ceramide, with a predominant content of sphingosine in mammals (d18:1). GlcCer is the precursor of a large scale of different glycosphingolipids. This cerebrozide is synthesized from uridine diphosphate-glucose and ceramide by a GlcCer synthase (UDP-glucose:ceramide glucosyltransferase; UGCG, EC 2.4.1.80). GlcCer-based sphingolipids have been identified as important mediators of a variety of cellular functions and their disequilibrium leads to pathological process development and may induce several diseases progression. Therefore, design of UGCG inhibitor represents an important topic for pharmaceutical research. In this paper, we aimed to study effects of newly synthesized derivatives of (±)-threo-1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (PPMP, known UGCG inhibitor) on: i) activity of UGCG in vitro; ii) thymocytes viability; iii) calcium transport through plasma membrane of thymocytes; iv) induction of apoptosis and autophagy in thymocytes. Thymocytes were isolated from thymus of three to seven weeks old mice (ICR strain). The key factors influencing the effect of PPMP analogues were their concentration, chemical structure and incubation time. Derivatives were able to change Ca2+ transport already after 15 min of cultivation, but their effects on cell viability were manifested at least after 12 h of cultivation. Four from fifteen studied compounds affected UGCG activity after four hour lasting cultivation, - but without correlation with data relating to effects on calcium transport and/or cell viability. Most potent UGCG inhibitor was chosen and applied for induction of apoptosis and autophagy in thymocytes. This inhibitor induced typical DNA fragmentation and upregulation of LC3B protein as autophagy marker, after 2 h and 4 h cultivation, respectively.

A COMPARISON OF THE SENSITIVITYOF DIFFERENT STRAINS OF MICE TO SARIN

Summary Poisoning from chemical warfare agents (CWAs) such as sarin is associated with neuronal degeneration. This damage is thought to result from glutamatergic excitotoxicity such as seen following kainic acid induced seizures. In order to search for novel neuroprotectants it is necessary to select good mouse models for susceptibility to nerve agent-induced seizures and the resulting neurodegeneration. The mouse strains tested (C57BL/6, ICR, DBA/2, SW, and FVB/N, Harlan Laboratory) had widely different sensitivity to sarin as shown by differences in the dose required resulting in 50% mortality, LD 50 . Differences also were observed among the strains in Fluoro-Jade C staining with the C57BL/6 and DBA having little to no staining when euthanized at 7 days whereas the other strains did. Differences in acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) activity were found among the strains as well. The ICR strain was excluded from the FOB and weight data due to difficulty getting a consistent LD 50 . Weight loss and FOB scores were similar for all strains. All strains had inhibited AChE activity after sarin exposure and exhibited inhibition of CNS BuChE after sarin exposure but only ICR and SW reached significance.

Pairmate-dependent pup retrieval as parental behavior in male mice.

Appropriate parental care by fathers can greatly facilitate healthy human family life. However, much less is known about paternal behavior in animals compared to those regarding maternal behavior. Previously, we reported that male ICR strain laboratory mice, although not spontaneously parental, can be induced to display maternal-like parental care (pup retrieval) when separated from their pups by signals from the pairmate dam (Liu et al., 2013). This parental behavior by the ICR sires, which are not genetically biparental, is novel and has been designated as pairmate-dependent paternal behavior. However, the factors critical for this paternal behavior are unclear. Here, we report that the pairmate-dependent paternal retrieval behavior is observed especially in the ICR strain and not in C57BL/6 or BALB/c mice. An ICR sire displays retrieval behavior only toward his biological pups. A sire co-housed with an unrelated non-pairing dam in a new environment, under which 38-kHz ultrasonic vocalizations are not detected, does not show parenting behavior. It is important for sires to establish their own home territory (cage) by continuous housing and testing to display retrieval behavior. These results indicated that the ICR sires display distinct paternity, including father-child social interaction, and shed light on parental behavior, although further analyses of paternal care at the neuroendocrinological and neurocircuitry levels are required.

Different behavioral effect dose–response profiles in mice exposed to two-carbon chlorinated hydrocarbons: Influence of structural and physical properties

The present study aimed to clarify whether dose–response profiles of acute behavioral effects of 1,2-dichloroethane (DCE), 1,1,1-trichloroethane (TCE), trichloroethylene (TRIC), and tetrachloroethylene (PERC) differ. A test battery involving 6 behavioral endpoints was applied to evaluate the effects of DCE, TCE, TRIC, and PERC in male ICR strain mice under the same experimental conditions. The behavioral effect dose–response profiles of these compounds differed. Regression analysis was used to evaluate the relationship between the dose–response profiles and structural and physical properties of the compounds. Dose–response profile differences correlated significantly with differences in specific structural and physical properties. These results suggest that differences in specific structural and physical properties of DCE, TCE, TRIC, and PERC are responsible for differences in behavioral effects that lead to a variety of dose–response profiles.

Angelica sinensis improves exercise performance and protects against physical fatigue in trained mice.

Angelica sinensis (AS) is a well-known medicinal herb and food material with antioxidative and multifunctional pharmacological activities. However, we lack evidence of the effect of AS on exercise performance and physical fatigue. We aimed to evaluate the potential beneficial effect of AS on ergogenic and anti-fatigue functions after physiological challenge. Male ICR strain mice were randomly assigned to four groups (n = 10 per group) for treatment: (1) sedentary control and vehicle treatment (vehicle control); (2) exercise training with vehicle treatment (exercise control); (3) exercise training with AS treatment at 0.41 g/kg/day (Ex-AS1); and (4) 2.05 g/kg/day (Ex-AS5); both the vehicle and AS were orally administered for 6 weeks. Exercise performance and anti-fatigue function were evaluated by forelimb grip strength, exhaustive swimming time, and levels of serum lactate, ammonia, glucose, and creatine kinase (CK) after a 15-min swimming exercise. Trend analysis revealed that AS treatments significantly increased endurance swimming time and blood glucose level, and decreased serum lactate, ammonia and CK levels. Liver and muscle glycogen contents were higher for Ex-AS1 and Ex-AS5 groups than the exercise control. Therefore, AS supplementation improved exercise performance and had anti-fatigue properties in mice and may be an effective ergogenic aid in exercise training.

Astragalus membranaceus improves exercise performance and ameliorates exercise-induced fatigue in trained mice.

Astragalus membranaceus (AM) is a popular “Qi-tonifying” herb with a long history of use as a Traditional Chinese Medicine with multiple biological functions. However, evidence for the effects of AM on exercise performance and physical fatigue is limited. We evaluated the potential beneficial effects of AM on ergogenic and anti-fatigue functions following physiological challenge. Male ICR strain mice were randomly assigned to four groups (n = 10 per group) for treatment: (1) sedentary control and vehicle treatment (vehicle control); (2) exercise training with vehicle treatment (exercise control); and (3) exercise training with AM treatment at 0.615 g/kg/day (Ex-AM1) or (4) 3.075 g/kg/day (Ex-AM5). Both the vehicle and AM were orally administered for 6 weeks. Exercise performance and anti-fatigue function were evaluated by forelimb grip strength, exhaustive swimming time, and levels of serum lactate, ammonia, glucose, and creatine kinase after 15-min swimming exercise. Exercise training combined with AM supplementation increased endurance exercise capacity and increased hepatic and muscle glycogen content. AM reduced exercise-induced accumulation of the byproducts blood lactate and ammonia with acute exercise challenge. Moreover, we found no deleterious effects from AM treatment. Therefore, AM supplementation improved exercise performance and had anti-fatigue effects in mice. It may be an effective ergogenic aid in exercise training.

Anticonvulsant profile of 2-ethylthio-7-methyl-4-(4-methylphenyl)pyrazolo[1,5-a][1,3,5]triazine

This work evaluates the central nervous effects in ICR strain mice of 2-ethylthio-7-methyl-4-(4-methylphenyl)pyrazolo[1,5-a][1,3,5]triazine (MH4b1), a compound obtained by an efficient one-step reaction of S,S-diethyl 4-methylbenzoylimidodithiocarbonate with 5-amino-3-methyl-1H-pyrazole, in order to assess its neuro-pharmacological profile. The tests applied were: maximal electroshock seizure (MES), pentylenetetrazole (PTZ) seizures, forced swimming, plus maze, marble burying, sleeping time, rota-rod and catalepsy. In addition, MH4b1 binding to the benzodiazepine site of the GABA-A receptor and MH4b1 inhibition of monoamine oxidase (MAO) subtypes A and B were evaluated. MH4b1 showed anticonvulsant effects in a dose dependent manner (30-300 mg/kg, p.o.) against MES and inhibition of MAO-B (IC50: 24.5 µM) without activity at the benzodiazepine site. These data suggest that MH4b1 has anticonvulsant properties related to MAO-B inhibition.

Differential Immune Response Associated to Malaria Outcome Is Detectable in Peripheral Blood following Plasmodium yoelii Infection in Mice

Malaria infection in humans elicits a wide range of immune responses that can be detected in peripheral blood, but we lack detailed long-term follow-up data on the primary and subsequent infections that lead to naturally acquired immunity. Studies on antimalarial immune responses in mice have been based on models yielding homogenous infection profiles. Here, we present a mouse model in which a heterogeneous course of Plasmodium yoelii lethal malaria infection is produced in a non-congenic ICR strain to allow comparison among different immunological and clinical outcomes. Three different disease courses were observed ranging from a fatal outcome, either early or late, to a self-resolved infection that conferred long-term immunity against re-infection. Qualitative and quantitative changes produced in leukocyte subpopulations and cytokine profiles detected in peripheral blood during the first week of infection revealed that monocytes, dendritic cells and immature B cells were the main cell subsets present in highly-parasitized mice dying in the first week after infection. Besides, CD4+CD25high T cells expanded at an earlier time point in early deceased mice than in surviving mice and expressed higher levels of intracellular Foxp3 protein. In contrast, survivors showed a limited increase of cytokines release and stable circulating innate cells. From the second week of infection, mice that would die or survive showed similar immune profiles, although CD4+CD25high T cells number increased earlier in mice with the worst prognosis. In surviving mice the expansion of activated circulating T cell and switched-class B cells with a long-term protective humoral response from the second infection week is remarkable. Our results demonstrate that the follow-up studies of immunological blood parameters during a malaria infection can offer information about the course of the pathological process and the immune response.

In Vitro Culture of Primary Testicular Stromal Cells derived from Mouse with Different Genetic Background : Optimization of Culture Temperature

Spermatogonial stem cells (SSCs) developed into sperms through spermatogenesis have been utilized as a useful tool in the field of regenerative medicine and infertility. However, a small number of highly qualified SSCs are resided in the seminiferous tubule of testis, resulted in developing effective in-vitro culture system of SSCs for solving simultaneously quantitative and qualitative problems. Presently, SSCs can be enriched on testicular stromal cells (TSCs), but there are no systematic researches about TSC culture. Therefore, we tried to optimize culture condition of TSCs derived from mouse with different strains. For these, proliferation and viability were measured and compared by culturing ICR outbred or DBA/2 inbred mouse-derived TSCs at 35 or 37℃. In case of ICR strain, primary TSCs cultured at 37℃ showed significantly higher proliferation and viability than those at 35℃ and significant increase of proliferation and viability in sub-passaged TSCs was detected in the 35℃ culture condition. Moreover, sub-passage of primary TSCs at 35℃ induced no significant effects on proliferation and viability. In contrast, in case of DBA/2 strain, significantly improved proliferation were detected in the primary TSCs cultured at 35℃, which showed no significant difference in the viability, compared to those at 37℃. Furthermore, sub-passaged TSCs cultured at 37℃ showed no significant differences in proliferation and viability, compared to those at 35℃. However, with significant decrease of proliferation induced by sub-passage of primary TSCs at 35℃, no significant effects on proliferation and viability were resulted from sub-passage of primary TSCs at 37℃. From these results, culture temperature of primary TSCs derived from outbred and inbred strain of mouse could be separately optimized in primary culture and subculture.

Enzymes That Control the Conversion ofL-Tryptophan-nicotinamide and the Urinary Excretion Ratio (N1-Methyl-2-pyridone-5-carboxamide +N1-Methyl-4-pyridone-3-carboxamide)/N1-Methylnicotinamide in Mice

There is little information on L-tryptophan→nicotinamide metabolism in mice. In the present study, we investigated the two important nutritional factors involved in metabolism L-tryptophan→nicotinamide; one is the amount of nicotinamide synthesized from L-tryptophan, and the other is the urine ratio (N(1)-methyl-2-pyridone-5-carboxamide + N(1)-methyl-4-pyridone-3-carboxamide)/N(1)-methylnicotinamide. The order of the percentages of nicotinamide synthesized from L-tryptophan was as follows: CBA strain mice (conversion percentage 0.41%) < BALB strain mice (0.82%) < C57BL/6 strain mice (1.13%) < ICR strain mice (1.70%). Urinary excretion of quinolinic acid was correlated with urinary excretion of the sum of nicotinamide and its catabolites (p<0.0001). The urine sum, which reflects the conversion of L-tryptophan→nicotinamide, correlated well with the activity of 3-hydroxyanthranilic acid dioxygenase (p=0.040). A nutritional indicator, the urine ratio (N(1)-methyl-2-pyridone-5-carboxamide + N(1)-methyl-4-pyridone-3-carboxamide)/N(1)-methylnicotinamide, was controlled by the activity of N(1)-methyl-2-pyridone-5-carboxamide-forming N(1)-methylnicotinamide oxidase.

CD38 in the nucleus accumbens and oxytocin are related to paternal behavior in mice

BackgroundMammalian sires participate in infant care. We previously demonstrated that sires of a strain of nonmonogamous laboratory mice initiate parental retrieval behavior in response to olfactory and auditory signals from the dam during isolation in a new environment. This behavior is rapidly lost in the absence of such signals when the sires are caged alone. The neural circuitry and hormones that control paternal behavior are not well-understood. CD38, a membrane glycoprotein, catalyzes synthesis of cyclic ADP-ribose and facilitates oxytocin (OT) secretion due to cyclic ADP-ribose-dependent increases in cytosolic free calcium concentrations in oxytocinergic neurons in the hypothalamus. In this paper, we studied CD38 in the nucleus accumbens (NAcc) and the role of OT on paternal pup retrieval behavior using CD38 knockout (CD38−/−) mice of the ICR strain.ResultsCD38−/− sires failed to retrieve when they were reunited with their pups after isolation together with the mate dams, but not with pup, in a novel cage for 10 min. CD38−/− sires treated with a single subcutaneous injection of OT exhibited recovery in the retrieval events when caged with CD38−/− dams treated with OT. We introduced human CD38 in the NAcc of CD38−/− sires using a lentiviral infection technique and examined the effects of local expression of CD38. Pairs of knockout dams treated with OT and sires expressing CD38 in the NAcc showed more retrieval (83% of wild-type sire levels). Complete recovery of retrieval was obtained in sires with the expression of CD38 in the NAcc in combination with OT administration. Other paternal behaviors, including pup grooming, crouching and huddling, were also more common in CD38−/− sires with CD38 expression in the NAcc compared with those in CD38−/− sires without CD38 expression in the NAcc.ConclusionsCD38 in the NAcc and OT are critical in paternal behavior.