NK Cells Require Cell-Extrinsic and -Intrinsic TYK2 for Full Functionality in Tumor Surveillance and Antibacterial Immunity.

Tyrosine kinase 2 (TYK2) is a widely expressed receptor-associated kinase that is involved in signaling by a variety of cytokines with important immune regulatory activities. Absence of TYK2 in mice results in impaired NK cell maturation and antitumor activity, although underlying mechanisms are largely unknown. Using conditional ablation of TYK2 in NK cells we show that TYK2 is required for IFN-γ production by NK cells in response to IL-12 and for an efficient immune defense against Listeria monocytogenes Deletion of TYK2 in NK cells did not impact NK cell maturation and IFN-γ production upon NK cell activating receptor (actR) stimulation. Similarly, NK cell-mediated tumor surveillance was unimpaired upon deletion of TYK2 in NK cells only. In line with the previously reported maturation-associated Ifng promoter demethylation, the less mature phenotype of Tyk2-/- NK cells correlated with an increased CpG methylation at the Ifng locus. Treatment with the DNA hypomethylating agent 5-aza-2-deoxycytidine restored the ability of Tyk2-/- NK cells to produce IFN-γ upon actR but not upon IL-12 stimulation. NK cell maturation was dependent on the presence of TYK2 in dendritic cells and could be rescued in Tyk2-deficient mice by treatment with exogenous IL-15/IL-15Rα complexes. IL-15 treatment also rescued the in vitro cytotoxicity defect and the impaired actR-induced IFN-γ production of Tyk2-/- NK cells. Collectively, our findings provide the first evidence, to our knowledge, for a key role of TYK2 in the host environment in promoting NK cell maturation and antitumor activity.

Aggressive B-cell lymphomas in patients with myelofibrosis receiving JAK1/2 inhibitor therapy.

Inhibition of Janus-kinase 1/2 (JAK1/2) is a mainstay to treat myeloproliferative neoplasms (MPN). Sporadic observations reported the co-incidence of B-cell non-Hodgkin lymphomas during treatment of MPN with JAK1/2 inhibitors. We assessed 626 patients with MPN, including 69 with myelofibrosis receiving JAK1/2 inhibitors for lymphoma development. B-cell lymphomas evolved in 4 (5.8%) of 69 patients receiving JAK1/2 inhibition compared with 2 (0.36%) of 557 with conventional treatment (16-fold increased risk). A similar 15-fold increase was observed in an independent cohort of 929 patients with MPN. Considering primary myelofibrosis only (N = 216), 3 lymphomas were observed in 31 inhibitor-treated patients (9.7%) vs 1 (0.54%) of 185 control patients. Lymphomas were of aggressive B-cell type, extranodal, or leukemic with high MYC expression in the absence of JAK2 V617F or other MPN-associated mutations. Median time from initiation of inhibitor therapy to lymphoma diagnosis was 25 months. Clonal immunoglobulin gene rearrangements were already detected in the bone marrow during myelofibrosis in 16.3% of patients. Lymphomas occurring during JAK1/2 inhibitor treatment were preceded by a preexisting B-cell clone in all 3 patients tested. Sequencing verified clonal identity in 2 patients. The effects of JAK1/2 inhibition were mirrored in Stat1-/- mice: 16 of 24 mice developed a spontaneous myeloid hyperplasia with the concomitant presence of aberrant B cells. Transplantations of bone marrow from diseased mice unmasked the outgrowth of a malignant B-cell clone evolving into aggressive B-cell leukemia-lymphoma. We conclude that JAK/STAT1 pathway inhibition in myelofibrosis is associated with an elevated frequency of aggressive B-cell lymphomas. Detection of a preexisting B-cell clone may identify individuals at risk.

High activation of STAT5A drives peripheral T-cell lymphoma and leukemia.

Recurrent gain-of-function mutations in the transcription factors STAT5A and much more in STAT5B were found in hematopoietic malignancies with the highest proportion in mature T- and natural killer-cell neoplasms (peripheral T-cell lymphoma, PTCL). No targeted therapy exists for these heterogeneous and often aggressive diseases. Given the shortage of models for PTCL, we mimicked graded STAT5A or STAT5B activity by expressing hyperactive Stat5a or STAT5B variants at low or high levels in the hematopoietic system of transgenic mice. Only mice with high activity levels developed a lethal disease resembling human PTCL. Neoplasia displayed massive expansion of CD8+ T cells and destructive organ infiltration. T cells were cytokine-hypersensitive with activated memory CD8+ T-lymphocyte characteristics. Histopathology and mRNA expression profiles revealed close correlation with distinct subtypes of PTCL. Pronounced STAT5 expression and activity in samples from patients with different subsets underline the relevance of JAK/STAT as a therapeutic target. JAK inhibitors or a selective STAT5 SH2 domain inhibitor induced cell death and ruxolitinib blocked T-cell neoplasia in vivo We conclude that enhanced STAT5A or STAT5B action both drive PTCL development, defining both STAT5 molecules as targets for therapeutic intervention.

Towards Establishing Cross-Platform Interoperability for Sensors in Smart Cities.

Typically, smart city projects involve complex distributed systems having multiple stakeholders and diverse applications. These applications involve a multitude of sensor and IoT platforms for managing different types of timeseries observations. In many scenarios, timeseries data is the result of specific simulations and is stored in databases and even simple files. To make well-informed decisions, it is essential to have a proper data integration strategy, which must allow working with heterogeneous data sources and platforms in interoperable ways. In this paper, we present a new lightweight web service called InterSensor Service allowing to simply connect to multiple IoT platforms, simulation specific data, databases, and simple files and retrieving their observations without worrying about data storage and the multitude of different APIs. The service encodes these observations "on-the-fly" according to the standardized external interfaces such as the OGC Sensor Observation Service and OGC SensorThings API. In this way, the heterogeneous observations can be analyzed and visualized in a unified way. The service can be deployed not only by the users to connect to different sources but also by providers and stakeholders to simply add further interfaces to their platforms realizing interoperability according to international standards. We have developed a Java-based implementation of the InterSensor Service, which is being offered free as open source software. The service is already being used in smart city projects and one application for the district Queen Elizabeth Olympic Park in London is shown in this paper.

The TSC-mTOR signaling pathway regulates the innate inflammatory response.

The innate inflammatory immune response must be tightly controlled to avoid damage to the host. Here, we showed that the tuberous sclerosis complex-mammalian target of rapamycin (TSC-mTOR) pathway regulated inflammatory responses after bacterial stimulation in monocytes, macrophages, and primary dendritic cells. Inhibition of mTOR by rapamycin promoted production of proinflammatory cytokines via the transcription factor NF-kappaB but blocked the release of interleukin-10 via the transcription factor STAT3. Conversely, deletion of TSC2, the key negative regulator of mTOR, diminished NF-kappaB but enhanced STAT3 activity and reversed this proinflammatory cytokine shift. Rapamycin-hyperactivated monocytes displayed a strong T helper 1 (Th1) cell- and Th17 cell-polarizing potency. Inhibition of mTOR in vivo regulated the inflammatory response and protected genetically susceptible mice against lethal Listeria monocytogenes infection. These data identify the TSC2-mTOR pathway as a key regulator of innate immune homeostasis with broad clinical implications for infectious and autoimmune diseases, vaccination, cancer, and transplantation.

Impact of superovulation and mating on the wellbeing of juvenile and adult C57BL/6N mice.

Superovulation of mice is routinely used to increase the number of obtainable ova per female. Because of the better outcome, prepubescent females are preferentially used. Here, we provide results of the impact of superovulation and mating on the wellbeing of juvenile compared with adult C57BL/6N mice. Two groups of mice (3-4 weeks vs 7-8 weeks old) were superovulated and mated. Observation of mating behaviour showed that reluctant adult females tended to fight the male's approach, whereas juveniles preferred to take flight. Faeces were collected daily for the analysis of stress hormones. There was no difference in the levels of glucocorticoid metabolites either between age groups or between treated animals and their controls. Histology after mating revealed intact vaginal mucosa without any detectable lesions in all animals regardless of age. In contrast to adults, almost all juveniles were synchronised in oestrus and produced significantly more ova. Taken together, our results reveal no increased welfare problem from using juvenile mice for superovulation and mating. Considering the higher yield of fertilisable oocytes and zygotes, it is advisable to use C57BL/6N prepubescent mice in order to reduce the number of donor females required.

A mouse model to identify cooperating signaling pathways in cancer.

We here establish a mouse cancer model called Multi-Hit that allows for the evaluation of oncogene cooperativities in tumor development. The model is based on the stochastic expression of oncogene combinations ('hits') that are mediated by Cre in a given tissue. Cells with cooperating hits are positively selected and give rise to tumors. We used this approach to evaluate the requirement of Ras downstream effector pathways in tumorigenesis.

Conditional ablation of TYK2 in immunity to viral infection and tumor surveillance.

Tyrosine kinase 2 (TYK2) has a pivotal role in immunity to infection and tumor surveillance. It is associated with several cytokine receptor chains including type I interferon (IFN) receptor 1 (IFNAR1), interleukin- (IL-) 12 receptor beta 1 (IL-12Rb1) and IL-10R2. We have generated a mouse with a conditional Tyk2 null allele and proved integrity of the conditional Tyk2 locus. TYK2 was successfully removed by the use of ubiquitous and tissue-specific Cre-expressing mouse strains. Myeloid TYK2 was found to critically contribute to the defense against murine cytomegalovirus. Ubiquitous TYK2 ablation severely impaired tumor immunosurveillance, while deletion in myeloid, dendritic or T cells alone showed no effect. The conditional Tyk2 mouse strain will be instrumental to further dissect TYK2 functions in infection, inflammation and cancer.

Growth-hormone-induced signal transducer and activator of transcription 5 signaling causes gigantism, inflammation, and premature death but protects mice from aggressive liver cancer.

UNLABELLED: Persistently high levels of growth hormone (GH) can cause liver cancer. GH activates multiple signal-transduction pathways, among them janus kinase (JAK) 2-signal transducer and activator of transcription (STAT) 5 (signal transducer and activator of transcription 5). Both hyperactivation and deletion of STAT5 in hepatocytes have been implicated in the development of hepatocellular carcinoma (HCC); nevertheless, the role of STAT5 in the development of HCC as a result of high GH levels remains enigmatic. Thus, we crossed a mouse model of gigantism and inflammatory liver cancer caused by hyperactivated GH signaling (GH(tg) ) to mice with hepatic deletion of STAT5 (STAT5(Δhep) ). Unlike GH(tg) mice, GH(tg) STAT5(Δhep) animals did not display gigantism. Moreover, the premature mortality, which was associated with chronic inflammation, as well as the pathologic alterations of hepatocytes observed in GH(tg) mice, were not observed in GH(tg) animals lacking STAT5. Strikingly, loss of hepatic STAT5 proteins led to enhanced HCC development in GH(tg) mice. Despite reduced chronic inflammation, GH(tg) STAT5(Δhep) mice displayed earlier and more advanced HCC than GH(tg) animals. This may be attributed to the combination of increased peripheral lipolysis, hepatic lipid synthesis, loss of hepatoprotective mediators accompanied by aberrant activation of tumor-promoting c-JUN and STAT3 signaling cascades, and accumulation of DNA damage secondary to loss of cell-cycle control. Thus, HCC was never observed in STAT5(Δhep) mice. CONCLUSION: As a result of their hepatoprotective functions, STAT5 proteins prevent progressive fatty liver disease and the formation of aggressive HCC in the setting of hyperactivated GH signaling. At the same time, they play a key role in controlling systemic inflammation and regulating organ and body size.

A novel Ncr1-Cre mouse reveals the essential role of STAT5 for NK-cell survival and development.

We generated a transgenic mouse line that expresses the Cre recombinase under the control of the Ncr1 (p46) promoter. Cre-mediated recombination was tightly restricted to natural killer (NK) cells, as revealed by crossing Ncr1-iCreTg mice to the eGFP-LSLTg reporter strain. Ncr1-iCreTg mice were further used to study NK cell-specific functions of Stat5 (signal transducers and activators of transcription 5) by generating Stat5(f/f) Ncr1-iCreTg animals. Stat5(f/f) Ncr1-iCreTg mice were largely devoid of NK cells in peripheral lymphoid organs. In the bone marrow, NK-cell maturation was abrogated at the NK cell-precursor stage. Moreover, we found that in vitro deletion of Stat5 in interleukin 2-expanded NK cells was incompatible with NK-cell viability. In vivo assays confirmed the complete abrogation of NK cell-mediated tumor control against B16F10-melanoma cells. In contrast, T cell-mediated tumor surveillance against MC38-adenocarcinoma cells was undisturbed. In summary, the results of our study show that STAT5 has a cell-intrinsic role in NK-cell development and that Ncr1-iCreTg mice are a powerful novel tool with which to study NK-cell development, biology, and function.

Comparison of unilateral and bilateral embryo transfer in mice.

Surgical embryo transfer in mice is a key technique in assisted reproduction and applied for different purposes in biomedical research. Due to its frequent application in rodent facilities across the world, further improvement of the procedure can substantially contribute to fulfil the principles of the 3Rs. Here, we investigated the effect of bilateral and unilateral left- or right-sided oviduct transfers on the success of embryo transfers. In total, we performed 223 embryo transfers (56 unilateral left, 56 unilateral right, 111 bilateral), in which we transferred 10-14 two-cell embryos each. We found that the type of transfer significantly influenced both the pregnancy rate of recipients and the survival rate of transferred embryos. Bilateral transfers yielded higher pregnancy and survival rates than left-sided unilateral transfers. Right-sided unilateral transfers yielded higher pregnancy rates than left-sided unilateral transfers and did not differ in embryo survival rates from bilateral transfers. We found no evidence that the number of transferred embryos affected the pregnancy rate. However, the number of born pups increased with the number of transferred embryos. In conclusion, unilateral embryo transfers into the right reproductive tract yield equally high pregnancy and embryo survival rates as bilateral transfers. Given that a second abdominal incision can be prevented and the time of surgery can be reduced, we recommend applying unilateral right-sided transfers, as this would reduce postoperative pain and lower the impact on recipients.

Inducible overexpression of a FAM3C/ILEI transgene has pleiotropic effects with shortened life span, liver fibrosis and anemia in mice.

FAM3C/ILEI is an important factor in epithelial-to-mesenchymal transition (EMT) induction, tumor progression and metastasis. Overexpressed in many cancers, elevated ILEI levels and secretion correlate with poor patient survival. Although ILEI's causative role in invasive tumor growth and metastasis has been demonstrated in several cellular tumor models, there are no available transgenic mice to study these effects in the context of a complex organism. Here, we describe the generation and initial characterization of a Tet-ON inducible Fam3c/ILEI transgenic mouse strain. We find that ubiquitous induction of ILEI overexpression (R26-ILEIind) at weaning age leads to a shortened lifespan, reduced body weight and microcytic hypochromic anemia. The anemia was reversible at a young age within a week upon withdrawal of ILEI induction. Vav1-driven overexpression of the ILEIind transgene in all hematopoietic cells (Vav-ILEIind) did not render mice anemic or lower overall fitness, demonstrating that no intrinsic mechanisms of erythroid development were dysregulated by ILEI and that hematopoietic ILEI hyperfunction did not contribute to death. Reduced serum iron levels of R26-ILEIind mice were indicative for a malfunction in iron uptake or homeostasis. Accordingly, the liver, the main organ of iron metabolism, was severely affected in moribund ILEI overexpressing mice: increased alanine transaminase and aspartate aminotransferase levels indicated liver dysfunction, the liver was reduced in size, showed increased apoptosis, reduced cellular iron content, and had a fibrotic phenotype. These data indicate that high ILEI expression in the liver might reduce hepatoprotection and induce liver fibrosis, which leads to liver dysfunction, disturbed iron metabolism and eventually to death. Overall, we show here that the novel Tet-ON inducible Fam3c/ILEI transgenic mouse strain allows tissue specific timely controlled overexpression of ILEI and thus, will serve as a versatile tool to model the effect of elevated ILEI expression in diverse tissue entities and disease conditions, including cancer.

Genetic background effects of keratin 8 and 18 in a DDC-induced hepatotoxicity and Mallory-Denk body formation mouse model.

Keratin 8 (K8) and keratin 18 (K18) form the major hepatocyte cytoskeleton. We investigated the impact of genetic loss of either K8 or K18 on liver homeostasis under toxic stress with the hypothesis that K8 and K18 exert different functions. krt8⁻/⁻ and krt18⁻/⁻ mice crossed into the same 129-ola genetic background were treated by acute and chronic administration of 3,5-diethoxy-carbonyl-1,4-dihydrocollidine (DDC). In acutely DDC-intoxicated mice, macrovesicular steatosis was more pronounced in krt8⁻/⁻ and krt18⁻/⁻ compared with wild-type (wt) animals. Mallory-Denk bodies (MDBs) appeared in krt18⁻/⁻ mice already at an early stage of intoxication in contrast to krt8⁻/⁻ mice that did not display MDB formation when fed with DDC. Keratin-deficient mice displayed significantly lower numbers of apoptotic hepatocytes than wt animals. krt8⁻/⁻, krt18⁻/⁻ and control mice displayed comparable cell proliferation rates. Chronically DDC-intoxicated krt18⁻/⁻ and wt mice showed a similarly increased degree of steatohepatitis with hepatocyte ballooning and MDB formation. In krt8⁻/⁻ mice, steatosis was less, ballooning, and MDBs were absent. krt18⁻/⁻ mice developed MDBs whereas krt8⁻/⁻ mice on the same genetic background did not, highlighting the significance of different structural properties of keratins. They are independent of the genetic background as an intrinsic factor. By contrast, toxicity effects may depend on the genetic background. krt8⁻/⁻ and krt18⁻/⁻ mice on the same genetic background show similar sensitivity to DDC intoxication and almost resemble wt animals regarding survival, degree of porphyria, liver-to-body weight ratio, serum bilirubin and liver enzyme levels. This stands in contrast to previous work where krt8⁻/⁻ and krt18⁻/⁻ mice on different genetic backgrounds were investigated.

Repeated use of surrogate mothers for embryo transfer in the mouse.

Embryo transfer in mice is a crucial technique for generation of transgenic animals, rederivation of contaminated lines, and revitalization of cryopreserved strains, and it is a key component of assisted reproduction techniques. It is common practice to use females only once as surrogate mothers. However, their reuse for a second embryo transfer could provide hygienic and economic advantages and conform to the concept of the 3Rs (replace, reduce, refine). This investigation evaluated the potential for a second embryo transfer in terms of feasibility, reproductive results, and experimental burden for the animal. Virgin female ICR mice (age 8-16 wk) were used as recipients for the first embryo transfer. Immediately after weaning of the first litter, a second surgical embryo transfer was performed into the same oviduct. Virgin females of comparable age to the reused mothers served as controls and underwent the same procedure. The first surgery did not affect the success of the second embryo transfer. Histological sections showed excellent wound healing without relevant impairment of involved tissues. We observed no differences in pregnancy rates or litter sizes between the transfer groups. Most importantly, we found no change in behavior indicating reduced well-being and no increase of corticosterone metabolites in the feces of surrogate mothers reused for a second embryo transfer. We conclude that a second embryo transfer in mice is feasible with regard to reproductive and animal welfare aspects.

Generation of mice with a conditional Stat1 null allele.

Interferons (IFNs) are key cytokines in the innate immune response that also bridge the gap to adaptive immunity. Signaling upon stimulation by IFN type I, II and III is mediated by the Jak-Stat pathway. STAT1 is activated by all three IFN receptor complexes and absence of STAT1 from mice increases their susceptibility to pathogens. In addition, depending on the setting, STAT1 can act as tumor suppressor or oncogene. Here we report the generation and detailed functional characterization of a conditional Stat1 knockout mouse. We show the integrity of the conditional Stat1 locus and report successful in vivo deletion by means of a ubiquitous and a tissue-specific Cre recombinase. The conditional Stat1 null allele represents an important tool for identifying novel and cell-autonomous STAT1 functions in infection and cancer.

Amniotic fluid stem cells and the cell source repertoire for non-invasive prenatal testing.

Cell-free fetal DNA (cffDNA)-based non-invasive prenatal testing (NIPT) is considered to be a very promising screening tool for pregnant women with an increased risk of fetal aneuploidy. Already millions of women worldwide underwent NIPT. However, due to the observed false-positive and false-negative results, this screening approach does not fulfil the criteria of a diagnostic test. Accordingly, positive results still require risk-carrying invasive prenatal testing, such as amniocentesis or chorionic villus sampling (CVS), for confirmation. Such hurdles need to be overcome before NIPT could become a diagnostic approach widely used in the general population. Here we discuss new evidence that besides the placenta amniotic fluid stem cells (AFSCs) could also represent an origin of cffDNA in the mother's blood. A comprehensive picture of the involved cell source repertoire could pave the way to more reliable interpretations of NIPT results and ameliorate counselling of advice-seeking patients.

Effect of Different Ambient Temperatures on Reproductive Outcome and Stress Level of Lactating Females in Two Mouse Strains.

Ambient temperature is an important non-biotic environmental factor influencing immunological and oncological parameters in laboratory mice. It is under discussion which temperature is more appropriate and whether the commonly used room temperature in rodent facilities of about 21 °C represents a chronic cold stress or the 30 °C of the thermoneutral zone constitutes heat stress for the animals. In this study, we selected the physiological challenging period of lactation to investigate the influence of a cage temperature of 20 °C, 25 °C, and 30 °C, respectively, on reproductive performance and stress hormone levels in two frequently used mouse strains. We found that B6D2F1 hybrid mothers weaned more pups compared to C57BL/6N mothers, and that the number of weaned pups was reduced when mothers of both strains were kept at 30 °C. Furthermore, at 30 °C, mothers and pups showed reduced body weight at weaning and offspring had longer tails. Despite pronounced temperature effects on reproductive parameters, we did not find any temperature effects on adrenocortical activity in breeding and control mice. Independent of the ambient temperature, however, we found that females raising pups showed elevated levels of faecal corticosterone metabolites (FCMs) compared to controls. Peak levels of stress hormone metabolites were measured around birth and during the third week of lactation. Our results provide no evidence of an advantage for keeping lactating mice in ambient temperatures near the thermoneutral zone. In contrast, we found that a 30 °C cage temperature during lactation reduced body mass in females and their offspring and declined female reproductive performance.

Dendritic cells require STAT-1 phosphorylated at its transactivating domain for the induction of peptide-specific CTL.

Phosphorylation of transcription factor STAT-1 on Y701 regulates subcellular localization whereas phosphorylation of the transactivating domain at S727 enhances transcriptional activity. In this study, we investigate the impact of STAT-1 and the importance of transactivating domain phosphorylation on the induction of peptide-specific CTL in presence of the TLR9-dependent immune adjuvant IC31. STAT-1 deficiency completely abolished CTL induction upon immunization, which was strongly reduced in animals carrying the mutation of the S727 phospho-acceptor site. A comparable reduction of CTL was found in mice lacking the type I IFN (IFN-I) receptor, whereas IFN-gamma-deficient mice behaved like wild-type controls. This finding suggests that S727-phosphorylated STAT-1 supports IFN-I-dependent induction of CTL. In adoptive transfer experiments, IFN-I- and S727-phosphorylated STAT-1 were critical for the activation and function of dendritic cells. Mice with a T cell-specific IFN-I receptor ablation did not show impaired CTL responses. Unlike the situation observed for CTL development S727-phosphorylated STAT-1 restrained proliferation of naive CD8(+) T cells both in vitro and following transfer into Rag-deficient mice. In summary, our data reveal a dual role of S727-phosphorylated STAT-1 for dendritic cell maturation as a prerequisite for the induction of CTL activity and for T cell autonomous control of activation-induced or homeostatic proliferation.

The transcriptional regulator PLZF induces the development of CD44 high memory phenotype T cells.

Transcriptional pathways controlling the development of CD44(hi) memory phenotype (MP) T cells with "innate-like" functions are not well understood. Here we show that the BTB (bric-a-brac, tramtrack, broad complex) domain-containing protein promyelocytic leukemia zinc finger (PLZF) is expressed in CD44(hi), but not in CD44(lo), CD4(+) T cells. Transgenic expression of PLZF during T cell development and in CD4(+) and CD8(+) T cells induced a T cell intrinsic program leading to an increase in peripheral CD44(hi) MP CD4(+) and CD8(+) T cells and a corresponding decrease of naïve CD44(lo) T cells. The MP CD4(+) and CD8(+) T cells produced IFNgamma upon PMA/ionomycin stimulation, thus showing innate-like function. Changes in the naïve versus memory-like subset distribution were already evident in single-positive thymocytes, indicating PLZF-induced T cell developmental alterations. In addition, CD1d-restricted natural killer T cells in PLZF transgenic mice showed impaired development and were severely reduced in the periphery. Finally, after anti-CD3/CD28 stimulation, CD4(+) transgenic T cells showed reduced IL-2 and IFNgamma production but increased IL-4 secretion as a result of enhanced IL-4 production of the CD44(hi)CD62L(+) subset. Our data indicate that PLZF is a novel regulator of the development of CD44(hi) MP T cells with a characteristic partial innate-like phenotype.

Cytoplasmic Transfer Methods for Studying the Segregation of Mitochondrial DNA in Mice.

Heteroplasmic mice represent a valuable tool to study the segregation of different mtDNA haplotypes (mtDNAs with differing alleles) in vivo against a defined nuclear background. We describe two methods for the creation of such models, differing in the resulting initial heteroplasmy levels: (a) transfer of ooplasm and (b) fusion of two blastomeres. These methods result in typical heteroplasmy of 5% and 50% donor mtDNA , respectively. The choice of method depends on the aim of the study. By means of breeding even 100% donor mtDNA can be reached within a few generations.