Although somatic cells can be reprogrammed to pluripotent stem cells (PSCs) with pure chemicals, authentic pluripotency of chemically induced pluripotent stem cells (CiPSCs) has never been achieved through tetraploid complementation assay. Spontaneous reprogramming of spermatogonial stem cells (SSCs) was another non-transgenic way to obtain PSCs, but this process lacks mechanistic explanation. Here, we reconstructed the trajectory of mouse SSC reprogramming and developed a five-chemical combination, boosting the reprogramming efficiency by nearly 80- to 100-folds. More importantly, chemical induced germline-derived PSCs (5C-gPSCs), but not gPSCs and chemical induced pluripotent stem cells, had authentic pluripotency, as determined by tetraploid complementation. Mechanistically, SSCs traversed through an inverted pathway of in vivo germ cell development, exhibiting the expression signatures and DNA methylation dynamics from spermatogonia to primordial germ cells and further to epiblasts. Besides, SSC-specific imprinting control regions switched from biallelic methylated states to monoallelic methylated states by imprinting demethylation and then re-methylation on one of the two alleles in 5C-gPSCs, which was apparently distinct with the imprinting reprogramming in vivo as DNA methylation simultaneously occurred on both alleles. Our work sheds light on the unique regulatory network underpinning SSC reprogramming, providing insights to understand generic mechanisms for cell-fate decision and epigenetic-related disorders in regenerative medicine.
Male ; Mice ; Animals ; Cellular Reprogramming/genetics* ; Tetraploidy ; Pluripotent Stem Cells/metabolism* ; Induced Pluripotent Stem Cells/metabolism* ; DNA Methylation ; Spermatogonia/metabolism* ; Germ Cells/metabolism*

Self-organized blastoids from extended pluripotent stem (EPS) cells possess enormous potential for investigating postimplantation embryo development and related diseases. However, the limited ability of postimplantation development of EPS-blastoids hinders its further application. In this study, single-cell transcriptomic analysis indicated that the "trophectoderm (TE)-like structure" of EPS-blastoids was primarily composed of primitive endoderm (PrE)-related cells instead of TE-related cells. We further identified PrE-like cells in EPS cell culture that contribute to the blastoid formation with TE-like structure. Inhibition of PrE cell differentiation by inhibiting MEK signaling or knockout of Gata6 in EPS cells markedly suppressed EPS-blastoid formation. Furthermore, we demonstrated that blastocyst-like structures reconstituted by combining the EPS-derived bilineage embryo-like structure (BLES) with either tetraploid embryos or tetraploid TE cells could implant normally and develop into live fetuses. In summary, our study reveals that TE improvement is critical for constructing a functional embryo using stem cells in vitro.
Pregnancy ; Female ; Animals ; Mice ; Tetraploidy ; Blastocyst ; Embryo, Mammalian ; Cell Differentiation ; Embryonic Development

OBJECTIVE: To explore the genetic and clinical characteristics of near-tetraploidy/tetraploidy karyotype (NT/T) in patients with myelodysplastic syndrome (MDS). METHODS: Cytogenetic findings of 1576 inpatients with primary MDS were retrospective analyzed, among which 9 were diagnosed with NT/T. Clinical data including gender, age, morphology, genetic feature and prognosis were analyzed. RESULTS: The prevalence of MDS patients with NT/T (NT/T-MDS) among all cases was 0.57%. Karyotyping analysis suggested that eight MDS patients had sole NT/T, while the remainder one had a complex karyotype. In addition to the typical morphology of MDS, NT/T-MDS had unique morphology including huge blast, double-nuclear cell and irregular nuclear membrane. One NT/T-MDS patient gave up therapy, and the remaining eight underwent the first course of treatment, albeit with poor prognosis. Only one patient had complete remission, one had partial remission, three had no remission; and three had converted to acute myeloid leukemia. CONCLUSION NT/T-MDS is rare and has unique morphology. Generally, NT/T-MDS patients have poor prognosis. However, NT/T cannot be simply classified as high-risk group, but with consideration whether they have affected particular chromosomal structures as well as other clinical data.
Humans ; Karyotype ; Leukemia, Myeloid, Acute/complications* ; Myelodysplastic Syndromes/pathology* ; Prognosis ; Retrospective Studies ; Tetraploidy

Parthenogenetic embryonic stem cells (pESCs) derived from bi-maternal genomes do not have competency of tetraploid complementation, due to lacking of paternal imprinting genes. To make pESCs possess fully development potentials and similar pluripotency to zygote-derived ESCs, we knocked out one allelic gene of the two essential maternal imprinting genes (H19 and IG) in their differentially methylated regions (DMR) via CRISPR/Cas9 system and obtained double knock out (DKO) pESCs. Maternal pESCs had similar morphology, expression levels of pluripotent makers and in vitro neural differentiation potentials to zygotes-derived ESCs. Besides that, DKO pESCs could contribute to full-term fetuses through tetraploid complementation, proving that they held fully development potentials. Derivation of DKO pESCs provided a type of major histocompatibility complex (MHC) matched pluripotent stem cells, which would benefit research in regenerative medicine.
Animals ; Embryonic Stem Cells ; Gene Knockout Techniques ; Genomic Imprinting ; Mice ; Parthenogenesis ; Pluripotent Stem Cells ; Regenerative Medicine ; Tetraploidy

OBJECTIVETo diagnose chromosomal abnormalities in amniotic fluid cells by combining karyotyping and single nucleotide polymorphism array (SNP-array) analysis, and to explore the application of SNP-array in routine clinical practice.

METHODSConventional G banding was used to karyotype a fetal amniotic fluid sample and the corresponding peripheral blood samples from the parents, followed by SNP-array analysis of the fetal genomic DNA from the amniotic fluid.

RESULTSThe karyotype of the amniocytes was 47, XX, +mar. The marker chromosome was further identified as psu idic (22) (q11.2) by SNP-array analysis, revealing tetraploidy of a 1.7 Mb fragment in 22q11.1-q11.2 interval that involves the critical region for Cat eye syndrome.

CONCLUSIONA rare chromosomal abnormality was identified by combining conventional G banding and SNP-array. The high resolution SNP-array could provide more detailed information for determining the origin of chromosomal abnormalities.

Adult ; Amniotic Fluid ; cytology ; Aneuploidy ; Chromosome Disorders ; genetics ; Chromosomes, Human, Pair 22 ; genetics ; Eye Abnormalities ; genetics ; Female ; Humans ; Isochromosomes ; Karyotyping ; Polymorphism, Single Nucleotide ; Pregnancy ; Tetraploidy

PURPOSE: Platycodon grandiflorum (a domestic diploid variety, DV-PG) has been used as a food and component of various traditional oriental medicines. Although DV-PG is known to have an anti-allergic effect, little is known about the beneficial health effects of the tetraploid ‘Etteum’ variety in the Platycodon grandiflorum (TV-PG), which is a recently developed variety. In this study, we investigated the effect of TV-PG on the rat basophilic leukemia mast cell (RBL-2H3)-mediated allergic response. METHODS: To examine the effects of TV-PG on the allergic response, RBL-2H3 cells were sensitized with dinitropheny (DNP)-immunoglobin E, treated with various concentrations of TV-PG, and challenged with DNP-human serum albumin. We estimated cell granulation by measuring the release of β-hexosaminidase and production of inflammatory mediators by ELISA. RESULTS: TV-PG had no effect on the proliferation or cytotoxicity of RBL-2H3 cells within the concentration range of 0 to 200 µg/mL. TV-PG inhibited degranulation of RBL-2H3 cells by antigen stimulation in a dose-dependent manner. TV-PG also suppressed the production of inflammatory cytokines and mediators such as interleukin-4, tumor necrosis factor-α, prostagladin E2, and leukotriene B4 in RBL-2H3 cells by antigen stimulation. CONCLUSION: These results indicate that TV-PG exhibits anti-allergic activity via inhibition of degranulation as well as suppression of inflammatory mediators and cytokine release. These findings suggest that TV-PG may have potential as a preventive and therapeutic agent for the treatment of various allergic diseases.
Animals ; Basophils ; Cytokines ; Diploidy ; Enzyme-Linked Immunosorbent Assay ; Functional Food ; Hypersensitivity ; Inflammation Mediators ; Interleukin-4 ; Leukemia ; Leukotriene B4 ; Mast Cells ; Medicine, East Asian Traditional ; Necrosis ; Platycodon* ; Rats ; Serum Albumin ; Tetraploidy*

In order to investigate the genetic basis of morphological variation of tetraploid plantlets of Atractylodes macrocephala, diploid plantlets were taken as experimental material, sterile filtration colchicine was used to soak 0.5-1.0 cm long buds. The difference between morphology and stomatal of diploid and tetraploid of A. macrocephala was compared, and genome polymorphism was explored by AFLP. The results showed that the buds dipped in 0.1% colchicine solution for 36 h was optimal conditions to induce tetraploid of A. macrocephala with induction rate of 32.0%. Morphological indexes such as leaf area index, leaf length and width, the density of stomas and the number of chloroplast of tetraploid were distinctly different from diploid. Four hundred and fifty-one bands ranging with 80-500 bp were amplified with 24 pairs of primers, the rate of polymorphism was 32.59%. These amplification sites of diploid were different from tetraploid of A. macrocephala, and the differences in morphology of them were reflected in the DNA polymorphism.
Amplified Fragment Length Polymorphism Analysis ; methods ; Atractylodes ; genetics ; Sequence Analysis, DNA ; Tetraploidy

OBJECTIVETo establish and optimize the rapid propagation system of Polygonum multiflorum, as well as explore method for induction and identification of autotetraploid.

METHODPropagation medium was optimized by orthogonal test. The buds were immersed in colchicine solution with different concentrations for different time to select induction conditions for autotetraploid of P. multiflorum.

RESULTThe most appropriate propagation medium was MS medium supplemented with 1.0 mg x L(-1) 6-BA, 0.3 mg x L(-1) NAA, and 0.4 mg x L(-1) PP333. That the buds were soaked in 0.2% colchicine solution for 30 h, or soaked in 0.3% colchicine solution for 18 h, was optimal condition to induce autopolyploid of P. multiflorum with induction rate as high as 16.7%.

CONCLUSIONRapid propagation of P. multiflorum could be achieved by tissue culture. Furthermore, colchicine was an effective inducer of polyploidy, and 25 tetraploid lines were obtained through chromosome identification. The experiment laid a foundation for the wild resource conservation, superior varieties breeding of P. multiflorum.

Chromosomes, Plant ; genetics ; Culture Media ; metabolism ; Polygonum ; genetics ; growth & development ; metabolism ; Tetraploidy ; Tissue Culture Techniques ; methods

No abstract available.
Acute Disease ; Aged ; Aged, 80 and over ; Bone Marrow Cells/cytology/metabolism/pathology ; Cell Lineage ; Humans ; Immunophenotyping ; Karyotyping ; Leukemia/*genetics/metabolism/pathology ; Male ; *Tetraploidy

OBJECTIVETo study the effect of gas-turbine green discoloring and drying processing method on the quality of various Lonicerae Japonicae Flos herbs.

METHODDIKMA DiamonsilTM-C18 column (4.6 mm x 250 mm, 5 microm) was adopted using HPLC Waters 1525 and eluted with acetonitrile and 0.1% phosphate acid as the mobile phase. The flow rate was 1.0 mL x min(-1) , the column temperature was 25 degrees C the detection wavelength was 355 nm.

RESULTAfter being processed by the gas-turbine green discoloring and drying method, tetraploid Lonicerae Japonicae Flos showed a green color. The contents of chlorogenic acid and galuteolin were 5.31% and 0.105% , both significantly higher by 18.0% and 32.1% than those of diploid Lonicerae Japonicae Flos processed by the same method. The content of chlorogenic acid in tetraploid Lonicerae Japonicae Flos processed the gas-turbine green discoloring and drying method were also remarkably higher than that of tetraploid and diploid Lonicerae Japonicae Flos processed by traditional processing method of natural drying.

CONCLUSIONThe gas-turbine green discoloring and drying processing method is a new-type drying method suitable for tetraploid Lonicerae Japonicae Flos. Under the condition of gas-turbine green discoloring and drying processing, tetraploid Lonicerae Japonicae Flos shows much higher quality than Lonicerae Japonicae Flos, suggesting that it is a good variety worth popularizing and applying.

Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; standards ; Flowers ; chemistry ; genetics ; Hot Temperature ; Lonicera ; chemistry ; genetics ; Quality Control ; Technology, Pharmaceutical ; methods ; Tetraploidy