Comparative global immune-related gene profiling of somatic cells, human pluripotent stem cells and their derivatives: implication for human lymphocyte proliferation.

Human pluripotent stem cells (hPSCs), including embryonic stem cells (ESCs) and induced PSCs (iPSCs), represent potentially unlimited cell sources for clinical applications. Previous studies have suggested that hPSCs may benefit from immune privilege and limited immunogenicity, as reflected by the reduced expression of major histocompatibility complex class-related molecules. Here we investigated the global immune-related gene expression profiles of human ESCs, hiPSCs and somatic cells and identified candidate immune-related genes that may alter their immunogenicity. The expression levels of global immune-related genes were determined by comparing undifferentiated and differentiated stem cells and three types of human somatic cells: dermal papilla cells, ovarian granulosa cells and foreskin fibroblast cells. We identified the differentially expressed genes CD24, GATA3, PROM1, THBS2, LY96, IFIT3, CXCR4, IL1R1, FGFR3, IDO1 and KDR, which overlapped with selected immune-related gene lists. In further analyses, mammalian target of rapamycin complex (mTOR) signaling was investigated in the differentiated stem cells following treatment with rapamycin and lentiviral transduction with specific short-hairpin RNAs. We found that the inhibition of mTOR signal pathways significantly downregulated the immunogenicity of differentiated stem cells. We also tested the immune responses induced in differentiated stem cells by mixed lymphocyte reactions. We found that CD24- and GATA3-deficient differentiated stem cells including neural lineage cells had limited abilities to activate human lymphocytes. By analyzing the transcriptome signature of immune-related genes, we observed a tendency of the hPSCs to differentiate toward an immune cell phenotype. Taken together, these data identify candidate immune-related genes that might constitute valuable targets for clinical applications

Toll-like receptor 2 gene polymorphisms, pulmonary tuberculosis, and natural killer cell counts

<p>Abstract</p> <p>Background</p> <p>To investigate whether the toll-like receptor 2 polymorphisms could influence susceptibility to pulmonary TB, its phenotypes, and blood lymphocyte subsets.</p> <p>Methods</p> <p>A total of 368 subjects, including 184 patients with pulmonary TB and 184 healthy controls, were examined for TLR2 polymorphisms over locus -100 (microsatellite guanine-thymine repeats), -16934 (T>A), -15607 (A>G), -196 to -174 (insertion>deletion), and 1350 (T>C). Eighty-six TB patients were examined to determine the peripheral blood lymphocyte subpopulations.</p> <p>Results</p> <p>We newly identified an association between the haplotype [A-G-(insertion)-T] and susceptibility to pulmonary TB (p = 0.006, false discovery rate q = 0.072). TB patients with systemic symptoms had a lower -196 to -174 deletion/deletion genotype frequency than those without systemic symptoms (5.7% vs. 17.7%; p = 0.01). TB patients with the deletion/deletion genotype had higher blood NK cell counts than those carrying the insertion allele (526 vs. 243.5 cells/μl, p = 0.009). TB patients with pleuritis had a higher 1350 CC genotype frequency than those without pleuritis (12.5% vs. 2.1%; p = 0.004). TB patients with the 1350 CC genotype had higher blood NK cell counts than those carrying the T allele (641 vs. 250 cells/μl, p = 0.004). TB patients carrying homozygous short alleles for GT repeats had higher blood NK cell counts than those carrying one or no short allele (641 vs. 250 cells/μl, p = 0.004).</p> <p>Conclusions</p> <p>TLR2 genetic polymorphisms influence susceptibility to pulmonary TB. TLR2 variants play a role in the development of TB phenotypes, probably by controlling the expansion of NK cells.</p

Tumor Spectrum, Tumor Latency and Tumor Incidence of the Pten-Deficient Mice

BACKGROUND: Pten functionally acts as a tumor suppressor gene. Lately, tissue-specific ablation of Pten gene in mice has elucidated the role of Pten in different tumor progression models. However, a temporally controlled Pten loss in all adult tissues to examine susceptibility of various tissues to Pten-deficient tumorigenesis has not been addressed yet. Our goal was to explore the genesis of Pten-deficient malignancies in multiple tissue lineages of the adult mouse. METHODS AND FINDINGS: We utilized an inducible Cre/loxP system to delete Pten exon 5 in the systemic organs of ROSA26 (R26)-CreER(T);Pten(fx/fx) mice. On reaching 45 weeks 4OHT-induced Pten loss, we found that the R26-CreER(T);Pten(fx/fx) mice developed a variety of malignancies. Overall tumor mean latency was 17 weeks in the Pten-deficient mice. Interestingly, mutant females developed malignancies more quickly at 10 approximately 11 weeks compared with a tumor latency of 21 weeks for mutant males. Lymphoma incidence (76.9% in females; 40.0% in males) was higher than the other malignancies found in the mutant mice. Mutant males developed prostate (20.0%), intestinal cancer (35.0%) and squamous cell carcinoma (10.0%), whereas the mutant females developed squamous cell carcinoma (15.4%) and endometrial cancer (46.1%) in addition to lymphomas. Furthermore, we tested the pharmacological inhibition of the PTEN downstream effectors using LY294002 on Pten-deficient prostate hyperplasia. Our data revealed that, indeed, the prostate hyperplasia resulting from the induced Pten loss was significantly suppressed by LY294002 (p = 0.007). CONCLUSIONS: Through monitoring a variety of Pten-deficient tumor formation, our results revealed that the lymphoid lineages and the epithelium of the prostate, endometrium, intestine and epidermis are highly susceptible to tumorigenesis after the Pten gene is excised. Therefore, this R26-CreER(T); Pten(fx/fx) mouse model may provide an entry point for understanding the role of Pten in the tumorigenesis of different organs and extend the search for potential therapeutic approaches to prevent Pten-deficient malignancies

A human in vitro model system for investigating genome-wide host responses to SARS coronavirus infection

10.1186/1471-2334-4-34BMC Infectious Diseases4-BIDM

A comparative study of the clock gene period in Blattella bisignata and B. germanica

日週律動現象普遍存在於生物當中，從原生動物到哺乳動物皆有一個近24小時的律動週期。果蠅的相關研究指出，生物時鐘的分子機制是由一個負回饋的機制所調控，時鐘細胞中period (per) 以及timeless (tim) 基因所產生的PER及TIM蛋白會抑制自身基因的表現。如同果蠅，其他昆蟲物種包括德國蜚蠊 (Blattella germanica)，per基因的表現具有日週律動的現象。在活動行為方面，德國蜚蠊的雄成蟲具有日週律動，但德國蜚蠊雌成蟲的活動行為受到生殖週期的影響，不具有日週律動的表現，此現象稱為掩蓋效應 (masking effect)。我們推測，生殖系統中的周邊時鐘 (peripheral clock)，會影響位於腦中的主要時鐘 (master clock) 對活動行為日週律動的表現。雙紋姬蠊 (B. bisignata) 是德國蜚蠊的近緣種，其雌蟲的活動行為日週律動表現不受生殖週期的影響，不具有掩蓋效應。本篇研究雙紋姬蠊與德國蜚蠊的時鐘基因per在頭部以及生殖系統中睪丸、卵巢與副腺的表現情況，試圖解釋掩蓋效應與日週律動的關係。選殖出的雙紋姬蠊per cDNA序列與德國蜚蠊per cDNA序列具有高度的相似性。根據Real-Time RT PCR的結果，per mRNA在雙紋姬蠊與德國蜚蠊的雄蟲頭部與部分生殖組織當中皆有相似的表現，其表現量在夜晚達到高峰。此外，per mRNA在雙紋姬蠊與德國蜚蠊雌蟲頭部與卵巢的表現顯示，per基因在這些組織當中似乎扮演另一種的調控角色，而與雄蟲頭部的表現情況不同。Circadian rhythms are nearly 24-hour rhythms of biological processes that persist under constant environmental conditions. These rhythms have been described in a variety of organisms ranging from prokaryotes to mammals. In Drosophila, the driving force underlying the circadian system is the pacemaker cells with time signal generated by the interactions of the negative feedback loop of period and timeless genes. In several other insect species, including the German cockroach, per expression undergoes circadian fluctuations. Oddly, the overt rhythm of locomotion of female German cockroach is masked by its own reproductive cycle. We suspect that clocks in peripheral tissues, especially in reproductive organs, may affect the brain-clock (master clock) which has been demonstrated to control the locomotor circadian rhythm. However, the Blattella bisignata, a closely-related species of the B. germanica, expresses locomotor circadian rhythm without masking effect in both sexes. This study focuses on the expression of per in brain and reproductive tissues in both species to unveil the possible interaction of clock gene expression under masking effect of locomotor circadian rhythm. Period gene in the double-striped cockroach has been cloned and its partial sequence shows a high degree of homology with the one from the German cockroach. Furthermore, the expression patterns of period mRNA were similar in heads and reproductive tissues, which were peaked at night. The exceptions in expression patterns of testes of German cockroach and ovaries of both species suggest that different regulatory clock mechanisms maybe involved.Table of Contents Literature review……………………………………………………………..1 Molecular clock……………………………………………………………1 Master and peripheral clocks…………………………………………………..5 Comparison between Blattella bisignata and B. germanica……………...10 Calling and mating behavior of the female German cockroach………...11 Reproductive system of the male Blattella germanica………………….12 Posttranslational regulation of PER……………………………………...14 Introduction…………………………………………………………………...17 Materials and Methods……………………………………………………….20 Insects………………………………………………………………………20 Reverse transcription (RT)-PCR………………………………………….20 Subcloning and sequencing………………………………………………..21 Rapid amplification of cDNA ends (RACE)……………………………...22 Computer analysis………………………………………………………….22 Real Time RT-PCR…………………………………………………………22 Western blotting…………………………………………………………….23 Results………………………………………………………………………….26 Cloning of period cDNA from Blattella bisgnata………………………..26 Sequence homology and structure analysis of PER………………………27 Temporal expression of period mRNA in heads and reproductive tissues of Blattella germanica…………………………………………………………29 Temporal expression of period mRNA in heads and reproductive tissues of Blattella bisignata…………………………………………………………..30 Temporal expression of PER in heads and reproductive tissues of Blattella germanica……………………………………………………………………32 Discussion………………………………………………………………………33 References………………………………………………………………………41 Appendix………………………………………………………………………..74 Acknowledgements…………………………………………………………….8

Bipolar Electrochemical Synthesis of WS 2

WS2 nanoparticles are prepared using bipolar electrochemistry. Obtained material exhibits high activity for hydrogen evolution reaction (HER) and it is used as a label in standard magneto-immunosandwich assay for protein detection through HER. This new system shows high analytical performance in terms of a wide range, selectivity, sensitivity, and reproducibility.NRF (Natl Research Foundation, S’pore)MOE (Min. of Education, S’pore

Far out-of-equilibrium spin populations trigger giant spin injection into atomically thin MoS2

Injecting spins from ferromagnetic metals into semiconductors efficiently is a crucial step towards the seamless integration of charge- and spin-information processing in a single device1,2. However, efficient spin injection into semiconductors has remained an elusive challenge even after almost three decades of major scientific effort3,4,5, due to, for example, the extremely low injection efficiencies originating from impedance mismatch1,2,5,6, or technological challenges originating from stability and the costs of the approaches7,8,9,10,11,12. We show here that, by utilizing the strongly out-of-equilibrium nature of subpicosecond spin-current pulses, we can obtain a massive spin transfer even across a bare ferromagnet/semiconductor interface. We demonstrate this by producing ultrashort spin-polarized current pulses in Co and injecting them into monolayer MoS2, a two-dimensional semiconductor. The MoS2 layer acts both as the receiver of the spin injection and as a selective converter of the spin current into a charge current, whose terahertz emission is then measured. Strikingly, we measure a giant spin current, orders of magnitude larger than typical injected spin-current densities using currently available techniques. Our result demonstrates that technologically relevant spin currents do not require the very strong excitations typically associated with femtosecond lasers. Rather, they can be driven by ultralow-intensity laser pulses, finally enabling ultrashort spin-current pulses to be a technologically viable information carrier for terahertz spintronics.NRF (Natl Research Foundation, S’pore)ASTAR (Agency for Sci., Tech. and Research, S’pore)MOE (Min. of Education, S’pore)Accepted versio

Membrane-bound carboxypeptidase E facilitates the entry of eosinophil cationic protein into neuroendocrine cells

ECP (eosinophil cationic protein) is a major component of eosinophil granule proteins, and is used as a clinical biomarker for asthma and allergic inflammatory disease. ECP has been implicated in damage to the cell membrane of many tissue types, but the mechanism is not well known. In the present study, mECP–eGFP–6H, a recombinant fusion protein containing mature ECP (mECP), enhanced green fluorescence protein (eGFP) and a His(6) tag (6H), has been expressed, purified and added to GH3 neuroendocrine cells to study the internalization ability of ECP. We found that mECP–eGFP–6H entered into GH3 neuroendocrine cells and inhibited the growth of the cells with an IC(50) of 0.8 μM. By yeast two-hybrid screening and immunoprecipitation, we have identified a specific protein–protein interaction between mECP and CPE (carboxypeptidase E), a well characterized metalloprotease. Further in vivo yeast two-hybrid screening has also revealed that residues 318–387 located in a region of unknown function in mature CPE are indispensable for association with mECP. In addition, the uptake of mECP–eGFP–6H is suppressed by dominant-negative expression of the recycling defect mutant pre-pro-HA–CPE(S471A,E472A) in GH3 cells, suggesting that the entry of mECP–eGFP–6H is associated with the recycling of CPE in GH3 cells. Taken together, we have demonstrated that CPE possesses a novel function to facilitate the entry of ECP to neuroendocrine cells, and such an endocytotic process allows the cytotoxic ECP to inhibit growth of the target cells